npc highlights 13 special | April 2011 PECIAL · Palma (UU). The PhD students can use their prize...

59th ASMS Conference on Mass Spectrometry, Denver, USA

Bessensap, Museon, The Hague, The Netherlands

5th European Summer School “Proteomic Basics”, Brixen/Bressanone, South Tirol, Italy

HUPO 2011, Geneva, Switzerland

Life Sciences Momentum 2011, The Netherlands

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netherlandscentre

npc highlights 13 special | April 2011

Programme overview and selected topics about Science, Bioinformatics, Valorisation, Research Hotels, Communications and Collaborations

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� | NPC Highlights 13 | April 2011

Contents

Preface

News headlines

Interview with the NPC Executive Board; Albert Heck, Bert Poolman, Hermen Overkleeft, Rob Liskamp and Werner Most

HighLights

10 Science; Geert Kops, Liesbeth Veenhoff, Arzu Umar and Shabaz Mohammed

14 Bioinformatics; Bas van Breukelen and Peter Horvatovich

16 Research Hotels; Maarten Altelaar and Gerco Angenent

19 Valorisation; Bas Nagelkerken, Bobby Florea, Huib Ovaa, Per Haberkant and Hans van Leeuwen

24 Communications; Martje Ebberink

Keynote lectures NPC Progress Meeting 2011; Matthias Mann, Lukas Käll, Roland Annan and Anne-Claude Gingras

EU supported projects

NPC Top Publications 2009-2010

NPC Theses 2009-2010

NPC at a glance

NPC Facts and Figures 2009-2010

NPC Network

About

The Netherlands Proteomics Centre (NPC) is a strategic collaboration of research groups from six

universities, four academic medical centres and several research institutes and biotech companies.

With a scientific programme addressing key areas of proteomics in several projects, and special-

ised ‘research hotels’, the NPC performs high-quality research and knowledge transfer in an

international context. The NPC is part of the Netherlands Genomics Initiative.

In NPC Highlights researchers present progress and results from NPC projects of the scientific pro-

gramme and the research hotels. NPC Highlights is published by the Netherlands Proteomics Centre.

Netherlands Proteomics Centre

Padualaan 8

NL 3584 CH Utrecht

t +31 30 253 4564

e [email protected]

w www.netherlandsproteomicscentre.nl

Editorial Board

Albert Heck, Scientific Director NPC

Werner Most, Managing Director NPC

Bert Poolman, member Executive Board NPC

Herman Overkleeft, member Executive Board NPC

Rob Liskamp, member Executive Board NPC

Martje Ebberink, Communications NPC

Authors Speical Issue Highlights

Astrid van de Graaf, Lilian Vermeer,

Bastienne Wentzel, Marga van Zundert

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Frontpage

Overview covers of 12 previous issues of NPC Highlights.

Coordination and editing

Marian van Opstal

Bèta Communicaties, The Hague

Lay-out

Frans Koeman

F.Koeman DTP-Services, Zoetermeer

Photography Speical Issue Highlights

Thijs Rooijmans, Henk Veenstra, Bas van Breukelen

Printing

Bestenzet BV, Zoetermeer

To subscribe to NPC Highlights please

send an e-mail with your full name, organisation

and address to [email protected]

© �011 Copyright Netherlands Proteomics Centre

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prefacenetherlandscentre

It is with great pleasure that we present this special issue of NPC Highlights to you. As we are about halfway through NPC’s second term, it is a good time to provide insight into the driving forces behind our programme and the results so far. It was quite a challenge to bring together all the various initiatives within this lively network and to present everything in sufficient detail. Therefore, rather than being exhaustive, we have tried to give you an overview and provide just a few examples of the different elements in our programme that address science, but also international collaborations, valorisation, education and knowledge transfer.

This update will also be part of NPC’s formal evaluation by an international review committee in April 2011, carried out on behalf of our funding body, the Netherlands Genomics Initiative, embedded in the Dutch Science Research Council (NWO). The NPC board looks forward to presenting and discussing the NPC programme and its results, and hopes that this evaluation will help to further continue and improve our activities.

We are happy with this opportunity to present the progress proteomics technologies and the NPC have made over the last few years. The steady production of scientific articles, which clearly show the increasing impact, documents this development. Through the NPC’s network, proteomics technologies have been applied to and further developed in partnerships with many leading research groups, making proteomics a crucial element of today’s research in the life sciences. Together with other technological disciplines, we are looking for ways to organise and continue these collaborations based on scientific expertise and to fully embed them in national and international networks.

Apart from the scientific arguments, the current pressure on public funding also necessitates coordinated efforts for enabling technologies such as proteomics to enhance innovation via translating the benefits efficiently to technology development and other fields of research. The NPC has a successful model in place for this, and the results illustrated here provide good arguments for continuing with our core activities.

I hope you enjoy reading this special issue of NPC Highlights.

Werner Most, NPC Managing Director

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NPC theme leader Shabaz Mohammed (UU) has been awarded a Vidi Grant from NWO for his research Decoding Cellular Communication. With the 800 K€ subsidy for a five year period Mohammed can start his own research line and build his own research group.Internal communication is vital for cells to perform their role. Stem cells can become any cell type if the right signals are gener-ated. This study aims to develop tools to decode the internal communication of stem cells during such a transition.

NWO TOP-GO for René Medema and Shabaz Mohammed

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International BioBusiness Summer School 2011

NPC Progress Meeting 2011

On 13 January the NPC Progress Meeting 2011 took place at the Media Plaza, Utrecht. Over 200 people from inside and outside the NPC participated. This year was particularly special as the NPC organised theme sessions in the morning, during which the scientific progress of the projects was discussed. In the afternoon four international keynote speakers shared their insights on the latest developments in proteomics and bioinformatics. Poster prizes were awarded to Harald Albers (NKI-AVL), Marit Terweij (NKI-AVL) and Serena Di Palma (UU). The PhD students can use their prize of 1 K€ for a journey abroad to attend a conference, summer school or other activity which is directly related to their research.

Antoni van Leeuwenhoek Award 2010 to Celia Berkers

On 17 January 2011, NPC researcher Celia Berkers (NKI-AVL) received the Antoni van Leeuwenhoek award 2010 for her research on various chemical tools to study the functioning of the proteasome with the aim to improve treatment with proteome inhibitors.The Antoni van Leeuwenhoek award is presented annually to a promising young researcher at the NKI-AVL. The prize consists of 6,5 K€ to be spent on studying or research. The prize money is provided by Roche Diagnostics Nederland BV.

Are you interested in pursuing your career in one of the many biomedical companies and you want to know more about the business field and your specific career opportunities? Then you should consider the BioBusiness Summer School, which will be held from 4-12 July in Amsterdam (AMC). The NPC will sponsor two NPC funded researchers to allow them to attend the BioBusiness Summer School. Researchers who would like to be considered for sponsoring can send their application to Martje Ebberink ([email protected]) up to 1 May 2011 at the latest. The application form can be downloaded from the NPC website.For more information about the summer school: www.biobusinesssummerschool.nl.

René Medema (left), UMCU, and NPC theme leader Shabaz Mohammed, UU, have been awarded a TOP-GO Grant by NWO of 750 K€ for their proposal Zicht op celdeling na DNA-schade (Insight into cell division after DNA damage).Widely used cancer therapies, such as radiation treatment and chemotherapy, kill cancer cells by damaging their DNA. However, despite the DNA damage, some cancer cells simply continue to divide, rendering therapy less effective. The researchers in this project systematically highlight the changes that occur when cell division starts again after the DNA damage. The new insights gained should ultimately lead to better cancer treatment.

Vidi awarded to Shabaz Mohammed

� | NPC Highlights Special | April 2011

As in previous years, the NPC will again sponsor two PhD students to allow them to attend this year’s European Summer School in Proteomic Basics. The summer

school will be held from 31 July – 6 August at Kloster Neustift in Brixen/Bressanone, South Tyrol, Italy. The subject this year is ‘From methods to clinical applications’. In addition to the standard 5-day programme, there will also be a clinical day.PhD students who would like to be considered for sponsoring can send their application to Martje Ebberink up to 15 April at the latest ([email protected]). The application form can be downloaded from the NPC website.For further information on the summer school: www.proteomic-basics.eu/

Fifth European ‘Proteomic Basics’ Summer School

Ernst Jung Medical Award 2011 for Hans Clevers

Hans Clevers, director of the Hubrecht Institute and NPC project leader, has been awarded the Ernst Jung Medical Award 2011, together with Christian Büchel (Hamburg University, Department of Systems Neuroscience). Clevers received this award in recognition of his pioneering research work in explaining the molecular causes of colon cancer which is expected to be of great benefit for the further development of innovative therapies.The award ceremony is scheduled for Friday 6 May 2010.

On Wednesday 8 December, Robbert Dijkgraaf presented the first Designers & Artists 4 Genomics Award (DA4GA) in the National Museum of Natural History Naturalis in Leiden. DA4GA — an initia-tive of the Waag Society, the Netherlands Genomics Initiative and the Centre for Society and

Genomics — is the first Dutch competition that brings together art, design and life sciences. According to the jury, the three winning designs are artistically and scientifically noteworthy and, furthermore, of social significance, thanks in part to the union of scientists and designers. The winners received 25 K€, which is to be used to complete their proposal that will then be on view in Naturalis from mid-June to late December 2011. For further information: www.society-genomics.nl

First Designers & Artists 4 Genomics Award presented

In a paper that appeared in Science, 4 February 2011, an inter-national consortium of scientists presents the genome sequence and organisation of Daphnia pulex, also known as the water flea. Jeroen Krijgsveld, former NPC theme leader and now team leader at EMBL Heidelberg, is a member of the Daphnia Genome Consortium, and contributed proteomics data to the study acquired when he was still in Utrecht.

Science publication: Genome organisation of the water flea

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Nature Chemistry publication: Unravelling formation of virus protection

The protective shell of a virus is made up of groups of proteins with the shape of a curved surface. This is the finding of NPC researcher Charlotte Uetrecht and her colleagues (UU). This fundamental research offers clues to block the formation of viruses or to turn the viruses into transporters of drugs. The results of the research are published in the scientific journal Nature Chemistry.Publication: Interrogating viral capsid assembly with ion mobility–mass spectrometry (Nature Chemistry 3, 126–132, 2011)

Scientific director of the NPC, Albert Heck, has been charged with coordi-nating PRIME-XS, a European consor-tium on proteomics. The European

Union has made 8 M€ available for this initiative. The partners of PRIME-XS, all leading in the field of proteomics, will provide access to their technology at six access facilities, distributed over Europe. In addition, the partners in the consortium devel-op new technologies to better aid the research community in answering current day scientific questions. The PRIME-XS kick-off event will take place on April 2 & 3 in Berlin, Germany. Project proposals can be submitted as from April the 4th. Calls for proposals are open for all scientists in EU mem-ber states and associated countries. Info: www.primexs.eu

Kick-off PRIME-XS

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The Netherlands Proteomics Centre is halfway its second period; a midterm review is at hand. An important moment, yet the board is looking further ahead, beyond 201�. “We want to stay in competition with the top proteomics groups,” stresses scientific director Albert Heck.

NPC’s five board members are not more than a healthy bit ner-vous about the coming midterm review. They have faith in a good outcome as the scientific output is substantial (77 articles in 2009 and 142 in 2010) and on international level. Scientific director Albert Heck: “I’m proud to present NPC’s work to the committee of international experts, and curious to hear their commentary.” Managing director Werner Most: “Although valo-risation requires time, we will also be able to present some

Looking past theproteomics hype

fine examples showing that our research is valuable in other re-search fields and to society.” He is referring to the phosphochip NPC developed together with Agilent Technologies. This lab-on-a-chip technology simplifies the analysis of phosphopeptides to a plug-and-go procedure. Another example is the discovery within the NPC that the proteins keratine 4 and cornuline are biomarkers for recurring head and neck cancer. Low quantities of these proteins at the incisions during tumour surgery are indicators that a tumour might recur. Now that it is possible to identify the high-risk patients, they can be monitored more intensively or treated preventively, resulting in a better prog-nosis. Board member Rob Liskamp also has good faith that NPC will receive good credits. Nevertheless, he is not too keen on the coming midterm review. He sighs: “We, Dutch scientists, are becoming the best writers of reports in the world. A bit more confidence from society as to how we spend our funds would be welcome these days. All these procedures take away a great deal of time from our actual work and passion: science.”

Firm groundsThe Netherlands Proteomics Centre, NPC, was established in 2003 with a grant of 24 Me from FES, a fund from the Ministry of Economics to strengthen the infrastructure in the Netherlands. The funding was received through the Netherlands Genomics Initiative, making the NPC part of the national Genomics strategy. Heck: “I vividly remember our visits to several judg-ing committees. It is a strange idea that politicians were re-quired to choose between a Genomics Initiative, including our proteomics centre, and a new highway around the city of Maastricht. Because at the end that was the decision they had to make.”

Interview

Bert Poolman“The atmosphere is amicable. Actually, we seldom disagree, but that is perhaps the result of NPC running so smoothly.”


important starting point for the development of new diagnostic tools and therapies for combating lysosomal storage disorders. Overkleeft: “The competition was close behind, but thanks to the NPC programme we succeeded in staying ahead and we now own a patent on the technology.” Werner Most adds that the ex-tra value of NPC also lies in providing the proteomics field with ‘a face’. “A centre is a visible and approachable entity. It connects experts and expertise. Just by forming a network, you identify the field and enlarge its potential.”

CommitmentBert Poolman, the fifth member on the board, characterises NPC as ‘unadorned’. Science and technology are the driving forces of the centre. There are no fancy trimmings: the overhead is low and effective to ensure that as much money as possible goes to equipment and researchers. Managing director Most emphasises that science has always been the leading factor in every choice; the organisational structure is accommodating. In spite of — or maybe thanks to — the lean administration, Overkleeft ranks NPC among the best managed institutes he is involved in. “I tend to think that science would be better off without managers, but directing over fifty know-it-all scientists

Genomics triumphed. And Heck became the scientific director of one of the world’s largest research centres in proteomics. The selection undoubtedly profited from proteomics being one of the newest buzz words in those days. “But I believe that the commitment of small biotech companies such as PamGene may well have been decisive,” says Liskamp. “NPC was not only backed up by the usual suspects such as DSM, Organon, Solvay and Philips, but also by a number of small, innovative start-up companies for whom this kind of financial commitment is significant.”Over the next four years NPC was able to spend a total of 62 Me on proteomics, as the FES budget was complemented with ac-ademic funds and large investments from industry. After the initial four years, NPC got good reviews and received an ad-ditional 22 Me for another four-and-a-half years. Heck: “I think we survived and passed the hype phase at that moment. NPC proved to provide added value and proteomics got recognition as a valuable technology.”As of mid 2013 the centre needs to find new sources of finan-cial support; at that time the FES funding will stop definitively. Board member Hermen Overkleeft: “Personally, I think it is a rather strange strategy that there is no prospect at all for con-tinuation of FES funding. FES initiated many research activities in highly potential fields; it would have been logical to award the most successful initiatives with a prospect of follow-up.” However, none of the board members sees 2013 as the final year of NPC. Liskamp: “When NPC started, proteomics was a kind of hype. Today, the field has developed enormously and gained firm ground. It has the potential to continue on its own strength.” Heck agrees that proteomics is now rooted and integrated in the Dutch scientific infrastructure, but he fears that without NPC proteomics in the Netherlands may not be able to keep up pace with the fast technology development worldwide. “Top scientists want to work with top technology, and in proteomics that means: expensive equipment. Proteomics research is a costly game to

play. Participating at the front line entails continuous invest-ments.” Overkleeft provides an example. At a Gordon confer-ence his colleague Hans Aerts presented their long sought-af-

ter discovery: a highly specific and sensitive activity-based probe with which the glycosidase that is deficient in Gaucher patients can be monitored in a chemical proteomics strategy. This is an

The NPC Board

• Albert Heck, professor of Biomolecular Mass Spectrometry, Utrecht University, scientific director

• Rob Liskamp, professor of Molecular Medicinal Chemistry, Utrecht University, board member

• Werner Most, NPC, managing director• Hermen Overkleeft, professor of Bioorganic Synthesis,

Leiden University, board member• Bert Poolman, professor of Biochemistry, University of

Groningen, board member

Albert Heck“Supported by a highly enthusiastic and motivated team, it is primarily a pleasure and an honour to be the chairman.”

We survived and passed the hype phase in those first four years

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isn’t an easy job. NPC handles it well, and, to be honest, with a minimum demand of my time and effort.” Another strong point of NPC Overkleeft mentions is that the research projects have the freedom to travel unexplored roads.

Making decisions about who you work with and who not is one of the most difficult tasks he has as the scientific director, Heck says. NPC gets many requests from scientists in the life sciences to support their research with proteomics techniques. NPC doesn’t have strict procedures, rules or guidelines to make these choices. “It just isn’t something that you can capture in a formal procedure,” Heck asserts. Each decision is made on the basis of a mix of the scientific reputation and commit-ment of the applicant, the nature of the research project and a kind of ‘gut feeling’. Heck: “Without a doubt, we have rejected some very interesting projects, but there is no better way.” One factor that has gained importance over the years is com-mitment. With state-of-the-art technology, instant success is rare. Therefore, a research partner needs to be as determined as NPC to invest time and energy in the project. Heck: “If they give up after one experiment, we have learned nothing. But I have also learned to ask: What if we succeed? What will be your next step? Our long term cooperations turn out to be the most successful. Therefore, we treasure our regular customers, but that unfortunately leaves little room for freshmen.”One lesson that Poolman learned in the first years of running his proteomics Research Hotel (the official name of the NPC col-laborative facilities) is that it is not efficient to let the lodgers do all the work themselves. “Being an academic, your initial approach is to teach visitors as much hands-on proteomics as possible. However, we soon realised that this is not efficient and also not necessary. Not all of our guests will use proteomics

technologies regularly, especially not the state-of-the-art equipment we use. They are better served by our scientists run-ning complex experiments for, or rather, with them. You don’t train a specialist within a few weeks.”

RewardingWhat has running a successful proteomics centre brought its scientific director? Heck: “One of the nicest aspects of the job is that I meet a lot of biologists, pharmaceutical and other scientists in the life sciences of high reputation, and learn about their scientific questions and problems. Sometimes I am jealous of these more conventional scientists. Some devote their en-tire career to one single protein or pathway. As a proteomics expert, I’m much more a generalist: I know a little of every hot topic in biology. This also makes me a kind of an outsider. Some academics still look at NPC as a kind of service centre. We are regarded as highly qualified technicians instead of top scientists. Yes, that grieves me a little at times, but being a generalist also has its great joys.”Poolman feels that he learns earlier about new scientific de-velopments thanks to his NPC board membership. Liskamp es-pecially enjoys the contacts with the non-scientists. “It’s quite interesting to see what captivates people from NGOs, politics and industry in what we do and in how we present it.” Most being the managing director of NPC neatly combines a challeng-ing number of his interests: science, public relations, manage-ment, finance, industry and policy.

Interview

Rob Liskamp“To be honest, my influence is modest. The course is set out by the directors. We act as the sounding board.”


And what has been the best NPC moment thus far? For Most a definite highlight was the 7th world congress of the Human Proteome Organisation (HUPO) that NPC hosted in Amsterdam in 2008. “First of all, it was an honour to be selected to organise such a great event. And when you note that everything is run-ning smoothly, that the presentations are fine, the discussions are lively, and the atmosphere among the 2000 participants is friendly and enjoyable, .... that’s just very rewarding.” The

scientists on the board have difficulty in naming a single best moment. Liskamp enjoys observing that proteomics finds its way into every pos-sible field of science, now

contributing for example to the development of personalised medicines. For Heck the joy is in many moments: in every top publication, in discussions with young scientists, and in starting up a brand new piece of equipment, and last but not least: the annual progress meetings attracting over 200 Dutch academic and industrial scientists interested in proteomics each year. “Not bad for such a small country,” he comments.

Beyond 201�NPC has just started its second period, yet the most discussed topic on the board at this time is NPC’s future after 2013, when FES funding ends. Scientific director Heck: “We all see that NPC adds value to the proteomics field in the Netherlands, therefore we are discussing various options for continuing.” Poolman: “Our discussions may seem premature to an outsider, but I can’t hold on to a good proteomics expert when I can’t guarantee him or her a steady job. Within a year, some will start looking for other job opportunities, therefore we need to open up new perspectives soon.”One option is to combine strengths with other ‘omics’ tech-nologies in a joint Dutch centre. A pilot project has already

started. Most: “Every technology must keep its scientific depth, but together we can integrate technological approaches and gain efficiency by combining administrative, communication and management tasks.”Another or additional strategy is attracting funds from various different sources. Most: “Several research groups now have a firm basis and a high scientific reputation. They may well suc-ceed in finding financers. Utrecht for instance has succeeded in attracting 8 Me in EU funding for PRIME-XS, a European consor-tium for protein research.” Heck agrees, but adds that living on various funds means writing a lot of applications and potentially many reviews. “But if it means NPC can continue, such an in-vestment is well spent.”Independent of what will come after 2013, Overkleeft con-cludes that NPC has raised proteomics in the Netherlands to a high level. Poolman: “A whole new generation of scientists is growing up with state-of-the-art proteomics. They fly out to industry and academic institutions here and abroad, creating a big network in advanced protein science. NPC has realised a head start. That alone creates opportunities in finding ways to continue the good work.”

Hermen Overkleeft“My role is foremost representing and defending the interests of the chemistry arm within NPC.”

Werner Most“The board members are very approachable. We meet about three times a year, but we often speak informally at conferences or over the phone.”

Possible future scenarios for the NPC

• Joining forces with other Dutch ‘omics’ technology centres – a pilot is running with the Netherlands Bioinformatics Centre, the Netherlands Metabolomics Centre, the Netherlands Consortium for Systems Biology, and the Centre for Society and Genomics

• A concise NPC based on various Dutch and/or European funds

• The strongest groups continue on the basis of their high scientific reputation and achievements and form a more informal network

Directing over fifty know-it-all scientists isn’t an easy job

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“Our main interest is the network of proteins that take care of cell division, and in particular mitosis. This is the phase in which the duplicated chromo-somes are separated and the cell divides into two daughter cells,” says Geert Kops. “A number of proteins regulate this process. We want to know when the regulatory proteins become active, and how they control the equal divi-sion of the chromosomes.” To elucidate these mechanisms, Kops also looks at tumour cells. “These cells make mistakes during cell divisions because their regulatory proteins do not efficiently see when a faulty separation is about to take place. The number of mistakes is limited; too many mistakes and the cell will die. If we understand how mitosis is regulated, we can try to intervene and even develop new anti-cancer treatments.”

Learn from each other“Most scientists are now aware that if you really want to understand this process, and many other processes in cell biology, you need proteomics. With proteomics one can get quantitative information about all the proteins that are present and how they ‘look’: their chemical modifications determine the function of the proteins in the cell. Examining RNA can only tell you so much. Modern research just can’t do without proteomics,” Kops concludes. “With proteomics you can also observe in great detail, follow a few proteins and their modifications in time, and observe how cell division is controlled.” Proteomics is a very complex and special technique. “To do it yourself demands a large investment in equipment and skills to operate such equipment. It is also something I don’t need continuously. Luckily, the NPC combines all these facilities and ex-pertise. We can make use of their expertise on mass spectrometry to answer our biological questions, and we can learn from each other,” says Kops. “This simply works the best. Especially since proteomics is a technique that also needs a lot of trouble shooting, which requires specialists to carry out.”

Next goalAs a theme leader, Kops organises a special theme day every year for all the project leaders to talk about their progress and to share ideas. There is also an annual session for NPC theme leaders to keep up with the developments. The past years were very successful for Kops. His group grew to ten mem-bers, working on eight projects. In addition, he was promoted to professor of molecular tumour cell biology in July 2010. His next goal is to become established as a top scientist in his field by maintaining high quality research over a long period of time. “All the research lines that have been set up now have the time to become really productive. And I have enough new ideas to go on.”

“Modern research can’t do without proteomics”Geert Kops is NPC theme leader for Cancer Proteomics and as of July 2010 full professor at the University Medical Centre Utrecht. In the coming years he is going to reap the harvest from all of his currently running projects on how cells divide properly.

“The NPC format is the perfect way to meet many other proteomics researchers”

Science

Grants• KWF Project 2010 (500 K€)• ERC Starting Independent Researcher

Grant 2009 (1.500 K€)• NWO Vidi Grant 2006 (600 K€)

Honours and Prizes• Elisabeth von Freyburgpenning, 2006

of the Catharijne Stichting• NVBMB Award, 2005 (Dutch society for

biochemistry and molecular biology)

Key publications• Jelluma, N. et al. (2010) Release of

Mps1 from kinetochores is crucial for timely anaphase onset. J Cell Biol 191, 281-290.

• Suijkerbuijk, S.J.E. et al. (2010) Molecular causes for BUBR1 dysfunc-tion in the human cancer predisposi-tion syndrome mosaic variegated aneuploidy. Cancer Res. 70, 4891-900.

• Jelluma, N. et al. (2008) Mps1 phos-phorylates Borealin to control Aurora B activity and chromosome alignment. Cell 132, 233-46.

In the spotlight

Geert KopsNPC theme leader ‘Cancer Proteomics’

University Medical Centre Utrecht

10 | NPC Highlights Special | April 2011

Liesbeth Veenhoff was working as a research fellow at the Rockefeller University in New York when her boyfriend, also working in New York, was of-fered a tenure track position at the University of Groningen. “We decided to go back to Groningen as this was an offer too good to refuse,” says Veenhoff. “I still had my fellowship money and would try to find a position. I wanted to continue working on the proteomics of the nuclear envelope.” Then Veenhoff

contacted Professor Bert Poolman, who had just become involved in the newly established Netherlands Proteomics Centre. “I could get funding for my work from the NPC. This was great because it allowed me to get started with my research and to learn a lot of

proteomics techniques. I optimised protocols for the reproducible isolation of pure and intact nuclear envelopes from yeast, the organism I use as a research model.”

A special ZIP codeRecently Veenhoff and co-workers investigated how membrane proteins (as opposed to non-membrane proteins) make it to the inside of the nucleus. It is known that the nuclear envelope contains nuclear pores, large pro-tein complexes that allow selective passage of molecules in and out of the nucleus. Veenhoff: “The big question is still: how are some of the proteins allowed inside the nucleus and others not.”The researchers discovered that membrane proteins contain a special ZIP code which indicates that their destination is the inner nuclear membrane. The ZIP code is located on a long linker which enables the binding via trans-port factors to the nuclear pore complex and the transport of the protein through the nuclear pore complex. “I am very excited about this,” says Veenhoff. “All the details being discovered on how the nuclear pore complex works, increase our understanding of what’s going on in this part of the cell and what and where it can go wrong. If nuclear transport doesn’t work, life is not possible.”

Inspiring to work for NPCVeenhoff is eager to unravel more details of the nuclear pore complex. In a future project she wants to find out which substances accumulate in the nucleus. Normally, during the isolation of nuclei, the contents leak out. “We now want to block the nuclear pore complexes so that the contents are pre-served and we can really find out what was there to begin with.”Veenhoff finds it inspiring to work for the NPC: “For many of my research questions I use proteomics methods. The NPC meetings are also usually very stimulating and sometimes lead to new collaborations and projects.”

Liesbeth VeenhoffNPC project leader ‘Yeast Nuclear Proteomics’

University Medical Centre Groningen

For many years Liesbeth Veenhoff has been studying the membrane that protects the nucleus in eukaryotic cells. Thanks to the NPC and other subsidies she was able to continue her research in unravelling the transport mechanisms in and out of the nucleus.

“I am especially interested in which proteins are inside the nuclear envelope”

Grants• NWO Vidi Grant 2008 (600 K€) • NWO Veni Grant 2004 (200 K€) • Fellowship Human Frontier Science

Programme 2002 (135 K€)

Honours and Prizes• Tenure Track UMC Groningen, 2010• Favourite cell biology paper by nature

editors, 2007• PhD, cum laude, 2001

Key publications• Meinema, A.C. et al. (2010) Long

unfolded linkers facilitate transport of membrane proteins through the central region of the nuclear pore complex, Submitted.

• Gauci, S. et al. (2009) Orthogonal sep-aration techniques for the character-ization of the yeast nuclear proteome. J Proteome Res 8(7), 3451-63.

• Alber, F. et al. (2007) The molecular architecture of the nuclear pore com-plex. Nature 450, 695-70.

In the spotlight

“I want to know how proteins end up on the inside of the

nucleus”

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Not everything goes as planned. “I was asked to speak at the Annual Meeting of the American Association for Cancer Research in April last year, a very important conference where twenty thousand scientists in cancer research gather,” says Arzu Umar. “That is a great honour. But then the volcano Eyjafjallajökull erupted and all flights were cancelled. I finally gave the presentation via a telephone call, and received wide applause.”

Turned to proteomicsThe first time Umar came in contact with proteomics was while she was working on her PhD on development of the male genital tract. “I was sup-posed to study RNA, but decided proteins were much more interesting and I turned to proteomics,” Umar affirms. After that she switched to research on breast cancer, a disease that affects one out of eight women in their life-time. “I thought if you want to achieve something in the scientific world and build your own research group, cancer research is the best field to be in.” Her KWF-fellow grant gave her the opportunity to gain experience abroad at the Pacific Northwest National Laboratory and to work on the new laser capture microdissection technique for finding new predictive biomarkers for hormonal therapy resistance. “A clump of tumour tissue is very heterogenic. Sometimes only 20 percent of the tissue consists of cancer cells. With laser microdissection we can cut out the tumour cells, collect them, and search very specifically for protein profiles.” When NPC was founded she worked closely with Theo Luider on his project, so she had time and space to develop herself. “To become a group leader, you need to bring in proj-ects. That means writing many project proposals and waiting for approval. After months of writing, it can take a year before approval is granted (or not). You must not be discouraged by setbacks.”

Loves academic researchIn 2009 Umar became a junior group leader. She now runs a group of four fellow workers. “One PhD student is working on a NPC project on prognostic factors for a special type of breast cancer called triple negative. This subtype is associated with a poor prognosis and lack of effective treatment. If we can find specific markers, this may finally lead to a better prediction of prognosis and develop-ment of new therapies. He is writing his first paper,” she says with pride.The one to one interaction with students is one of the reasons she loves aca-demic research best. “It is important to see my students grow and blossom, and I can help them do so by achieving something myself too and by working on my dream to be well-known worldwide for innovative research. I want to reach for the highest goal possible. NPC gave me the opportunity to show more of myself at my best.”

“Reach for the highest goal possible”Her dream is to become internationally recognised for outstanding research. The first steps are in place. Arzu Umar has built her own research group. One of her projects is for the NPC.

“NPC gave me the opportunity to show more of

myself at my best”

Science

Arzu UmarNPC project leader ‘Proteomics at breast cancer’

Erasmus Medical Centre Rotterdam

Grants• KWF Grant, 2009-2013 (535 K€) • Pink Ribbon Grant, 2009-2010 (125 K€)

Honours and Prizes• Speaker at Major Symposia on

Proteomics, AACR Annual Meeting, 2010• Scholar-In-Training award - AACR

Annual Meeting, 2008• KWF Fellowship, 2004-2006

Key publication• Umar, A. et al. (2009) Identification of

a putative protein-profile associating with tamoxifen therapy-resistance in breast cancer. Molecular & Cellular Proteomics 8 (6),1278-1294.

In the spotlight


After obtaining his PhD in 2003 at the University of Manchester, UK, Shabaz Mohammed went to Denmark to work with Professor Ole Jensen at the pro-tein research group. There he focused on developing new methodologies for proteomics. “Increasingly mass spectrometry was recognised as a powerful tool for answering all kinds of important biological questions, whereas when I was working on my PhD, we were still trying to understand the fundamen-tals of the technology.”

Methods of choiceMohammed continued to work on proteomics techniques for the NPC in the Department of Biomolecular Mass Spectrometry and

Proteomics at the Utrecht University. He pursued a number of research interests. Together with colleagues he developed a strategy for de novo sequencing of peptides. Mass spectrometry (MS) is the method of choice for identifying proteins, but it requires prior knowledge about the genome of the relevant species. Mohammed: “In the de novo sequencing method, this prior knowledge is no longer a prerequisite, allowing for a more flexible approach to identification. The principle ideas for the strategy rest on the choice of a ‘new’ enzyme and MS sequencing technique, ETD. The generated MS spectra are simpler and easier for sequence determination.” Phosphorylation is an important posttranslational modification of proteins. It can operate as an on/off switch for activity of a protein in vivo. However, phosphorylated proteins are usually present in lower concentrations than the unphosphorylated proteins, which makes their analysis in proteomics chal-lenging. Mohammed and colleagues developed a chip together with Agilent Technologies to enrich for phosphopeptides prior to MS analysis. There is a titanium dioxide (TiO2) trap in this chip, which aids enrichment of phospho-peptides. “The beauty of the chip is that it requires little knowledge to oper-ate, thus enabling biologists to analyse phosphorylation themselves,” says Mohammed. He also developed the application of hydrophilic interaction liquid chromatography (HILIC) for performing comprehensive analyses of cel-lular protein content. This separation method is proving to be an attractive choice for reducing the complexity of a protein sample prior to mass spectro-metric analysis since its demands on material levels are very modest.

Solving biological questions“I like to develop new methods with which researchers can solve their biological questions a lot faster,” says Mohammed. “It allows them to focus more on their main research question.” “The NPC has provided me the foundation for where I am now and the possi-bility to take it to a new level. It allowed me to generate many new ideas and offered the possibility of bringing these ideas to practice in an efficient way.”

Shabaz MohammedNPC theme leader ‘New Separation and Enrichment

Tools in Proteomics’

Utrecht University

Six years ago when Albert Heck asked Shabaz Mohammed to come to work for the NPC, Mohammed did not hesitate long. “After hearing about the possibilities and the resources, I jumped on board. It was a great opportunity for me to try to do something completely new.”

“The multifaceted environment of the NPC is highly rewarding”

Grants• NWO Vidi Grant, 2010 (800 K€)• NWO TOP-GO Grant, 2010 (750 K€,

together with René Medema)• ‘Nieuw Amsterdams Peil’ Award, 2008

(5 K€)

Honours and Prizes• Agilent Technologies Foundation

Research Awards• HUPO Young Investigator Prize, 2008

Key publications• Taouatas, N. et al. (2008) Straight

forward ladder sequencing of peptides by a combination of Lys-N proteolytic cleavage and electron transfer disso-ciation. Nature Methods 5(5), 405-7.

• Pinkse, M.W. et al. (2008) Highly ro-bust, automated, and sensitive online TiO2-based phosphoproteomics ap-plied to study endogenous phosphory-lation in Drosophila melanogaster. J Proteome Res 7, 687-97.

• Boersema, P.J. et al. (2008) Hydrophilic interaction liquid chroma-tography (HILIC) in proteomics. Anal Bioanal Chem 391(1), 151-9.

In the spotlight

“Collaborating with excellent biologists who ask fundamental

questions is very inspiring”

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The aim of the Gaining Momentum initiative, which was started in 2007 by the NPC together with the Netherlands Bioinformatics Centre (NBIC), is to

create a high quality workflow for the analysis of proteomics

data. “For some time now, the progression in proteomics has been hampered by what we call the data processing bottleneck,” explains Peter Horvatovich, who together with Bas van Breukelen leads the initiative. Both combine their chemistry and biochemistry education with ample experience

Peter Horvatovich (Analytical Biochemistry, University of Groningen) is theme leader of ‘Bioinformatics in Proteomics’.

Connecting bioinformatics and proteomicsGenerally, the ever increasing flow of data from proteomics experiments has led to a big demand on bioinformatics to develop the necessary infrastructure for the storage, processing, analysis and visualisation of proteomics data. Three years ago the ‘Gaining Momentum’ initiative was started to integrate bioinformatics in proteomics research. The theme leaders Bas van Breukelen and Peter Horvatovich assess the results so far.

in programming and bioinformatics. It is an ideal combination for the task of leading the Gaining Momentum initiative, which needs a global outlook to connect people from both sides.

Closing the gap“People in wet labs such as NPC researchers are producing more and more complex data using advanced technologies,” says Horvatovich. Most of these researchers are using very basic bioinformatics methods to analyse their data. They have neither the time nor the knowledge to learn, develop and test new software. On the other hand, he says, bioinformatics software developers such as the people working within NBIC are experts at writing code but have limited knowledge of the problems within proteomics research. “This is exactly the gap we want to close,” Horvatovich explains. “We are experts on both.”The high quality workflows are built in practice from a set of bioinformatics tools. Based on the workflows developed within the Gaining Momentum initiative, the goal is to provide user friendly, high-throughput data processing services to analyse proteomics LC-MS data. The programmes are developed by members of the platforms or are obtained from open source code repositories and must apply to a set of rules defined by the Gaining Momentum initiative to ensure that the various parts are modular and can be combined. “There is a tendency to reinvent the wheel again and again,” observes Bas van Breukelen. He explains: “By setting the standard for data representation following, for example, the HUPO Proteomics

Bioinformatics

Bas van Breukelen (Biomolecular Mass Spectrometry and

Proteomics Group, Utrecht University)

is the NPC hotel manager of the Bioinformatics

Research Hotel and coordinator of the

NPC-NBIC programme.

1� | NPC Highlights Special | April 2011

Software repository Trac is used as a software repository system. In this reposi-tory you can develop your programmes and collaborate with other NBIC-BioAssist programmers. The default versioning sys-tem is Subversion for new projects, but it is possible to use CVS for existing projects (http://trac.nbic.nl).

Gaining Momentum InitiativeTogether with the Netherlands Bioinformatics Centre (NBIC), the NPC has joined forces to set up a large task force: Gaining Momentum at the Proteomics-Bioinformatic Interface. This task force has the goals of building a platform for proteomics based on bioinformatics and of providing tools, workflows and high-throughput data process-ing services for the proteomics community.

Netherlands Bioinformatics for Proteomics PlatformNetherlands Bioinformatics for Proteomics Platform (NBPP, http://nbpp.nl) is a joint initiative of NBIC and the NPC. The goal of NBPP is to provide user friendly, high-throughput data processing services to analyse proteomics LC-MS data, based on open source tools or tools developed by and available to the Platform members. These services are provided by Data Analysis Framework using the Dutch Life Science Grid and other computational clusters of the platform members.

Galaxy serverThe NBIC Galaxy server is a development server where the implementation of new tools is tested. Hence, this is not a production server! The Galaxy server is maintained by the Netherlands Bioinformatics Centre (NBIC). For more info please visit: http://galaxy.nbic.nl.

Protein data processing services

Standards Initiative, we made sure that data is described in a uniform manner. It is now possible to connect the software building blocks so people can build their own workflow by combining modules written by others. Achieving this technical uniformity was very challenging indeed.”

Full suite of powerful softwareThe second important aspect is the documentation and visual interface that the programmes and workflows must have when published on the tool repository or when made available online as easy-to-use data processing services. Without proper documentation it is almost impossible for others to use the software. “The academic goal in bioinformatics is not only to develop the code and publish it, but also to make it available to others via data processing services that are easy-to-use. Otherwise no one will use it. This is exactly what our platform is doing,” says Horvatovich.The NBIC software repository is used to place the source code of the developed tools. The NBIC Galaxy server and the web page of the Netherlands Bioinformatics for Proteomics Platform (NBPP) are used to make online data processing

services available. The first workflows have recently been put online. “This is just the beginning. The ultimate goal is to provide a full suite of powerful software,” van Breukelen says. “But on a small scale, things are happening. For example, the StatQuant program, which also has a graphical user interface, has been available from the NBIC repository for some time. We have more than a thousand users already, but this programme also draws people to the repository to download other pro-grams.”

Ensuring continuationThe cooperation between NBIC and NPC has always been very good, both project leaders agree. Their main concern now is continuation of funding. One of the strategies to ensure continuation is to connect with other initiatives, says Horvatovich. “For example metabolomics can use the tools we are developing,” he explains. Van Breukelen agrees: “We are set up with a solid core. It has been hard to find the right people to work on the projects. It would be a shame to waste the efforts. It is our task now to show that proteomics cannot live without us.”

Bioinformatics in ProteomicsThe ever increasing flow of data from proteomics experiments have led to a large demand on bioinformat-ics to develop the necessary infrastructure for the storage, processing, analysis and visu-alisation of proteomics data. The NPC Enabling Technology Program ‘Bioinformatics in Proteomics’ (E4) especially focuses on proteomics driven bioinformatics. The E4 pro-gramme aspires to cover as many proteomics bioinformat-ics topics as possible with a maximum cross connectivity between the NPC research themes.

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Research Hotels

Hosts and guests benefit from NPC Research HotelsThe NPC offers the newest proteomics technologies and facilities in specialised Research Hotels for joint projects with external scientists. The host-guest formula has proven very helpful in achieving high level proteomics research and challenging cooperations. The success exceeds the capacity.

Gerco Angenent, hotel manager Analytical Hotel Wageningen:“The CBSG is likely to adopt our Research Hotel formula for its metabolomics and sequencing facilities”

Maarten Altelaar, hotel manager Analytical Hotel Utrecht:“Project proposals are set up in collaboration between the visiting group and the hotel manager”

“Proteomics analysis requires a lot of time and effort,” says hotel manager Maarten Altelaar. “That is why the NPC Research Hotels, with the

disposal of thorough proteomics skills and advanced pro-teomics technologies, meet the needs of many researchers.” Groups with expertise in biology but lacking time, funding or expertise in proteomics may seek assistance from one of the NPC Research Hotels via the NPC website or personal contacts. Ideally, the research is conducted in close cooperation, as opposed to analytical service laboratories that merely execute the requested analyses and send the data back.

Quality leadsAltelaar judges the project applications for quality, feasibil-ity and accordance with the research of his group. “We would like to accept all requests, but we simply do not have enough capacity to do that,” clarifies Altelaar. His Analytical Hotel Utrecht has facilities including a range of proteomics tech-niques such as several state-of-the-art mass spectrometry and liquid chromatography methods. The expertise of the group ranges from single protein identification and quantification to the study of interaction of protein complexes and posttransla-tional modifications in whole cell lysates. Hotel manager Gerco Angenent also judges projects for qual-ity and feasibility. “We make sure that samples and other preparations are made in the labs of the requesting groups. Here, we focus on MS analysis and, more importantly, on data interpretation. That is the major bottleneck in research.”

Unique selling pointAngenent’s Analytical Hotel in Wageningen is based both in the Plant Research Institute (PRI) and Wageningen University. Together they are specialised in mass spectrometry for unrav-elling signalling pathways and identifying protein complexes in plants. “The combination of proteomics facilities and bio-logical background is the unique selling point for the hotel.” Angenent organised his hotel facility under the NPCII in a man-ner different from most others. His group is, like many other Dutch plant research groups, strongly linked to the Centre for Biosystems Genomics (CBSG) in Wageningen. CBSG was highly interested in the hotel formula and decided to participate. Both NPC and CBSG contribute to the hotel. Unlike most of the


Hosts and guests benefit from NPC Research Hotels

Groningen

Utrecht

Wageningen

Rotterdam

Delft

Cell Sorting Hotel UtrechtCell specific proteomics. Equipment and ex-pertise to isolate specific cell types from plant tissues, to extract proteins from sorted cells and to subject them to proteomics analysis. (Utrecht University)

other hotels, the Wageningen Hotel gives priority to proj-ect propositions from within NPC or CBSG. “We do not have enough facilities to help everyone,” says Angenent, agreeing with Altelaar. “In this way, we make sure that the money, knowledge and expertise benefit the proteomics community in the Netherlands.” One of the projects that has gained momentum using the hotel facilities is the isolation of transcription factor complexes in plants. “Plant research is behind human or yeast research in this respect, mainly because we cannot use cell lines,” says Angenent. “Almost no transcription factor complexes have been isolated or identified. At the Wageningen Hotel we have isolated the first complexes and we will publish these shortly.”

Added value“The projects within the hotel facilities also have added value for us,” says Altelaar. “We are able to cooperate in projects that are outside the scope of our biological capabilities.”

Analytical Hotel UtrechtExpert centre for state of the art proteomics analysis. Innovative mass spectrometry methods are developed and implemented for the characterisation of biomol-ecules in their relation to their biological function.(Utrecht University)

Analytical Hotel RotterdamEmbedded within the Erasmus MC Proteomics Centre. Mass spectrome-try based proteomics methodologies and protein separation techniques for the qualitative and quantitative analysis of (sub)proteomes. (Erasmus Medical Centre Rotterdam)

Analytical Hotel GroningenPurification, separation, and high resolution analysis of proteins em-ploying advanced proteomics tools and techniques for membrane/ organelle isolation, ultracentrifu-gation and chromatography-based protein separation coupled to mass spectrometry identification.(University of Groningen)

Analytical Hotel WageningenLinked to activities within the Centre for Biosystems Genomics (CBSG). Focus on deciphering the genetic basis of plant specific processes, improving plant per-formance and increasing product quality.

(Plant Research International / Wageningen University)

Analytical Hotel DelftUp to date peptidomics to study native, endogenous peptides with important biological (regulatory, signal, (neuro)hormonal) functions. Focus on various peptide chem-ical identification strategies, including de novo sequencing. (Delft University of Technology)

Bioinformatics Hotel UtrechtDevelopment of bioinformatics software and algorithms that facilitate proteomics research including (raw) data (pre) process-ing, data storage and dissemination and knowledge extraction.(Utrecht University)

Cell Culture Hotel UtrechtKey technologies for targeted proteomics analyses of stem cells and differenti-ated cells. This includes selective tag-ging of proteins using bacterial artificial chromosomes (BACs) and viral-mediated gene manipulation. (University Medical Centre Utrecht)

An illustrative example is cooperation with cell biologists from the Netherlands Cancer Institute (NKI-AVL). They are interested in the formation of filopodia on migrating cells. Filopodia are the microspikes extending from migrating cells and are believed to be involved in sensing of their direct environment and directed locomotion of the cells. By studying the proteome of these filopodia, fundamental knowledge is gained which may lead to more understanding about tumour cell proliferation. “This cooperation, like many others initi-ated through the NPC hotels, is not a one-off matter,” explains Altelaar. “Many projects are in a second or third phase and will be pursued. The field of proteomics is continuously evolv-ing, giving others the opportunity to answer questions that were previously hard to tackle.”Neither Altelaar nor Angenent see major drawbacks in the hotel formula, except for the limited capacity to accept all applications. “Hotel guests are extremely enthusiastic,” they affirm, “and the hotel setting is greeted with interest.”

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I n d u s t r i e s

Proposals are selected for:• scientific quality• fit within the goals of the NPC• fit within the expertise of the respective NPC Hotel• feasibility within the indicated project time• quality of the visiting group

NPCResearch

Hotels

Du

tch

resea rch inst itutes and

univ

ers

itie

s

FO

RE

IGN

RESEARCH INSTITUTES ANDUNIV

ERSI

TIE

S

Join the expertsThe NPC offers the newest proteomics technologies and facilities in specialised Research Hotels for joint projects with external scientists. You can address your subject of interest in collaboration with experienced personnel in our Hotels. This figure

Industries

Foreign research institutes and universities

Dutch research institutes and universities

12

30

171

Total projects 2009-2010

32RUG

Groningen 23TUDDelft 21

UUUtrecht

20WUR

Wageningen

20UMCU

Utrecht

13EMC

Rotterdam

8HubrechtInstituteUtrecht

7NKI-AVL

Amsterdam

7PRI

Wageningen4

LULeiden

3UMCG

Groningen

3RUN

Nijmegen

3UvA

Amsterdam

1LUMCLeiden

1UM

Maastricht

1NIOO-KNAWWageningen

1NIZOEde

1UMCN

Nijmegen1

RIVMBilthoven

1VU

Amsterdam

2EMBL

Germany

2MPI

MartinsriedGermany

1Gene

CentreMunichGermany 1

Germany 2K.U.

LeuvenBelgium

2Uppsala

UniversitySweden

1KarlolinskaInstituteSweden

1CincinnatiChildren's

USA

1Cornell

UniversityUSA

1UC

BerkeleyUSA

1NIH

BethsedaUSA

1SUNY

Upstate USA

1Imperial College

UK

1QUBUK

1Unidad deBiofisicaSpain

1UAMSpain

1ETH

ZuerichSwitzerland

1IEOItaly

1FlemingInstituteGreece

1IGBMCFrance

1Academia

SinicaTaiwan

1IIMCBPoland 1

BTISingapore

1Universityof AlbertaCanada

Merus

AgilentTechno-logies

Profibrix Bioceros

Philips

Eli Lilly

Genmab

PamGene

Thermo Scientific

BrukerDaltonics

FEIElectron

Optics

Number of projects

BrukerNonius

UH Munster

HU Berlin 1

Germany

1UGentBelgium

shows the current national and international project partners (2009 - 2010).

Research Hotels

Dear NPC Highlights reader,

In this special issue of NPC Highlights I would like to take the opportunity to update you on some of the events from the past two years.

In 2009, the NPC started to implement a valorisation pro-gramme that supports those that have created an interesting invention and are looking for ways to move forward with their technologies. The NPC realised the urgent need for additional funding to properly validate a business opportunity. For this, we introduced the NPC Valorisation Voucher and many of you have already found your way through our application procedure.

The NPC also realised that valorisation is not part of standard education at many Dutch universities and institutes. Although most universities/medical centres by now have a decent tech-nology transfer office in place with skilled people to support the valorisation process, it is still a challenging process making researchers aware of what to expect when considering fil-ing a patent application for an invention and bringing that to market, for example. The NPC recognised this and decided to organise a Valorisation and Patent Workshop for people inside and outside the NPC network. The first workshop was held in November 2010 and attracted a small audience with an attractive programme covering the basics on the patent system, but also highlighting the relevance of patents for the industry and for starters. The audience greatly appreciated the latter in particular as they learned what a fellow researcher encoun-tered on his way to setting up a business venture.

The NPC also decided to make the valorisation programme part of the annual NPC Progress Meeting in order to monitor the progress of granted valorisation projects. During the 2011 NPC Progress Meeting all four project teams presented a status update to a small panel of experts in the tech transfer, industry and start-up fields. Lively discussions were held with relevant recommendations and suggestions for the teams. The NPC will certainly continue to have these meetings in the future.

So, what can we expect from the second half of NPC II? The NPC will continue to organise the NPC Valorisation and Patent workshop. We encourage you to make use of this service; after all, we organise it for you! We will also be looking at other ways to create awareness of valo-risation, perhaps in closer collaboration with some of the other NGI centres. Needless to say, we will continue to evaluate the NPC Valorisation Voucher applications as long as budget al-lows. We anticipate being able to fund an additional 5 projects. However, it’ll be first come, first served, so don’t hesitate if you have an opportunity that needs funding!

All the best,Bas NagelkerkenNPC Valorisation Manager

Netherlands Proteomics Centre

Contact+31 30 253 4564

[email protected]

Bas Nagelkerken

NPC Valorisation Manager

Valorisation

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Valorisation

A boutique for chemical probes

Repetitive requests to purchase compounds designed, synthesised and published by the bio-organic synthesis group in Leiden drove the scientists into a new metier. They professionalised their service and started their own supply company: Chemical Proteomics Reagents (CPR).

result of publications. But we are professionalising and we are going to do more marketing.”

Full package of servicesFlorea proudly mentions that for a year now supplier Enzo Life Sciences has had seven of the CPR compounds listed in the compound catalogues on their website. “Over the coming years we hope to make a deal with Enzo to offer twenty to thirty of our compounds. For that privilege we would like a structural annual payment. It will give us a solid basis for growth and for more investment in IP protection,” explains Florea. “The second phase is of a low risk character. At that time we will not only supply compounds for research, but hope also to be able for offering a full package of services and doing research for the industry as well.”

Milestones 2009 commercialised several (<10) products2009 set up an organisation for CPR activities 2009 NPC Valorisation Voucher 2010 professionalised the administration system2010 first large scale production2010 business plan2011... -further exploring the IP landscape

-extension commercial production (about 20 new compounds)-extension partnerships for sales and marketing -apply for the STW valorisation Grant

“It started out quite simply,” says biochemist Bobby Florea at the Leiden Institute for Chemistry. “As organic chemists we try to synthese organic molecules

with a biological function or application. After publication we receive many requests from the scientific community for a few micrograms of our compounds. “We used to honour these requests and would sometimes get a co-authorship offered in return. However, we became aware that potential users would rather have no strings attached and pay for the reagents.” That was the early start of what is now Chemical Proteomics Reagents (CPR). Florea formed a business team with Rian van den Nieuwendijk and Hermen Overkleeft.

Small specialised shop“We call it a boutique,” says Florea, following a sugges-tion from Kenn Curt Daniël of Leiden University Research &

Innovation Services, which facilitates the commercialisation of research in Leiden. “We adopted that idea. A boutique is a small specialised shop where you can buy stuff that is exclu-sive and the price is too,” laughs Florea. The NPC Valorisation Voucher gave Florea the opportunity to make the crucial steps towards a start-up company. He was able to work for some eighteen months on the business plan and to map the freedom to operate in the present IP landscape. In addition, Van den Nieuwendijk was able to test production on a larger scale. “Some research groups need 100 micrograms or up to a few grams of a compound, which is sometimes more than we have on the shelf.”The beginning is high risk according to Florea’s plan. “We start with the compounds and the activity-based probes we already have and acquire a market share first. Patents are too expen-sive, since our current annual profit is around ten thousand euro. This is without any marketing efforts and solely as a

Bobby Florea

“We are professionalising and we are going to do more marketing”20 | NPC Highlights Special | April 2011

Valorisation

UbiQ jumps in expanding ubiquitin research field

The synthesis of ubiquitin conjugates, biochemically speaking a mission impossible, was solved by Farid El Oualid and Huib Ovaa with organic chemistry. This finding was the stepping stone for launching their start-up company UbiQ. Now Ovaa is seeking a suitable lab location.

founded his company UbiQ along with postdoc Farid El Oualid and biotech entrepreneur Alfred Nijkerk. “We have a mixed business model,” Ovaa explains. “Right now we earn our money with the sales of reagents and assays. Boston Biochem, the world’s leading producer of ubiquitin-related products, offers our compounds for research and drug discovery. But we have our own ideas, too,” Ovaa smiles. “We are investing our profits in our next ambition: drug discovery.”

Next milestoneTo produce ubiquitin derivatives and conjugates in sufficient quantities in the near future, UbiQ has to move to a larger scale production and an independent lab facility. Therefore the entrepeneur is seeking a suitable lab location for his company. “It is really difficult. Lab spaces for start-ups in the life sciences are not easy to find but for now UbiQ has space within my lab at the NKI.”

Ubiquitin is a small regulatory protein which acts as posttranslational modifier that controls protein breakdown. If protein degradation is not con-

trolled correctly, disease could be the consequence. “You can compare its importance with the posttranslational phosphory-lation system,” affirms organic chemist Huib Ovaa, head of a Chemical Biology group at the Division of Cell Biology at the Netherlands Cancer Institute (NKI-AVL). “The role of phosphorylation is well understood, since the phosphorylated peptides for investigating the enzymes involved, like kinases or phosphatases, are available.”The posttranslational modification with ubiquitin is even more important in controlling cell processes, according to Ovaa. However, ubiquitin is a complex molecule and therefore very difficult to synthesise. And the enzymatic cascade that at-taches ubiquitin is not as simple as for phosphorylation. The conjugation of ubiquitin requires the concerted action of E1, E2 and E3 enzymes in defined combinations. This complexity has hampered research to present. “It is just impossible to make ubiquinated proteins using biochemical methods, with very few exceptions to this rule. But it is not impossible with innovative chemistry,” explains Ovaa, who figured out their synthesis together with postdoc Farid El Oualid. They decided to give it a try. “The first time we tested it, the reaction ran smoothly. It was so much more than we expected. Now we are able to basically synthesise anything needed for ubiquitin-re-lated research. And we are the first and only company in the world that can do this by fully automated synthesis.”

Basis to start offWith the support of the NKI, Ovaa applied for a patent on the chemical ligation technology. The Pre-Seed Grant of the NGI and the NPC Valorisation Voucher he received in 2009 provided a solid basis for starting off. Ovaa and El Oualid automated their developed technology to synthesise ubiquitinated peptides and conjugates, enabling the development of novel and powerful screening-assays. The NPC Valorisation Voucher was used to expand the product library with ubiquitin-like proteins, such as synthetic Nedd8 and SUMO conjugates, in order to expand options to target other drug targets, and therefore to obtain more market potential. A year later Ovaa

“We are investing our profits in our next ambition: drug discovery”

Milestones 2007 first idea for synthesis route of ubiquitin proteins 2008 first successful test 2009 NGI Pre-Seed Grant 2009 NPC Valorisation Voucher 2010 launch UbiQ company (www.ubiqbio.com)2010 international patent application (WO 2010131962 20101118)2011… search for lab location

Huib Ovaa

| 21

At the time Per Haberkant was studying a protein-lipid interaction with a photoactivatable lipid precursor during his PhD in Heidelberg, he had never

thought of click chemistry. It was in the section Membrane Enzymology at Utrecht University, which he joined as a post-doc, when he got acquainted with this approach and got his luminous idea. He figured out a new strategy to study protein-lipid interaction based on bi-functional lipid precursors that contain a photoactivatable group and a clickable moiety. “At first I could only look at one individual protein-lipid interac-tion. In combination with the clickable group, I can identify a whole class of proteins at the same time,” says Haberkant. “With this technology we can perform high-throughput screening on lipid-protein interactions. This is unique and has never been described before.” With support from the Utrecht University Holding BV, he filed a patent application.

Market potentialThe technology will give new insights into many fundamental research questions on the role of lipid-regulated processes in the cell. For instance, how lipids are involved in numerous signalling events, and how disturbed homeostasis of lipids con-tributes to diseases such as cancer, diabetes and Alzheimer’s disease. “To demonstrate the power of the technology, we identified the protein that interacts with ceramide, a putative tumour suppressor lipid. When this protein is inhibited cancer cells are more susceptible to chemotherapy and radiotherapy. The ceramide transfer protein is a target for further drug development.”Haberkant used the NPC Valorisation Voucher to develop new precursors to extend the application possibilities of the tech-nology. “We are designing novel probes to address the interac-tion of proteins with major lipid classes. In the meantime we are going to validate the technology by showing that a newly identified lipid-protein interaction is really taking place. This

Milestones 2008 publication on photoactivatable sphingosine for the in vivo analysis

of protein-sphingolipid interactions (J Lipid Res, 49, 251).2009 review, on the idea of combining photo activation with click

chemistry (Biol Chem, 390, 795).2009 US Patent Application (20110020837)2010 NPC Valorisation Voucher 2010 proof-of-concept2010 validation of newly-identified lipid protein interactions2011… bringing technology to the market

The combination of photocrosslinking with click chemistry formed the basis of Per Haberkant’s ingenious invention. The NPC Valorisation Voucher made it possible to deliver a proof-of-concept and to validate his new tool for large scale profiling of lipid-protein interactions.

New lipid-protein interaction tool in sight

Valorisation

is important to further commercialise the technology. Then we can bring it to the market.”

Business partnersAfter making his discovery, the inevitable question popped up: do you want to start your own company? “I thought about it briefly and decided not to. I am afraid that if I start a business based on this new tool, my time will be absorbed by things like finance and sales. But we can sell the technology to other companies. We are in discussion with large lipid producers like Avanti Polar Lipids and Echelon Biosciences, and they are very interested.” With the revenues Haberkant creates financial space for more fundamental research. That is what he loves the most.

“Business discussions for bringing our technology to the market are well under way”

Per Haberkant


Options for commercialisationUltimately this will lead to the development of a diagnostic tool for identifying increased risk of fracture at an early stage and for suggesting preventive treatment. “There are a few options for commercialising the identified biomarkers. Either we do it ourselves or we leave it to the diagnostic in-dustries,” says Van Leeuwen. “We have our own biotech com-pany, Arcarios BV, which focuses on bone anabolic drugs, but it is also possible we will sell it. Our research lab has built up good contacts with the diagnostic industry. The new biomark-ers would mean an important new step in the diagnostics of bone diseases and a great market opportunity.”

Novel biomarkers predicting bone fracture risk

Valorisation

To tackle the huge problem of increasing osteoporosis and bone frac-tures in the near future, there is a need for additional biomarkers for early detection and treatment. Luckily, Hans van Leeuwen is on the trail of finding new blood-based biomarkers for new diagnostic tools.

“The best predictor of osteoporosis and facture risk today is measuring the bone mineral density. People with low bone density have a high risk

of fracture,” according to Hans van Leeuwen, professor of calcium and bone metabolism at Erasmus Medical Centre in Rotterdam. However, this is only one part of the story. Studies on people with fractures in relation to bone mineral density (BMD) revealed surprising results: 50 to 70 percent of the people with fractures didn’t have osteoporosis according to the BMD criteria of the World Health Organisation. “This means that there are other factors that determine fracture risk, such as bone quality, i.e. the bone renewal capacity and the composition of the bone protein matrix.”

Targeted protein assaysSince bone is continuously renewed, the chance is high that proteins involved in bone formation or degradation and proteins in the bone matrix can be found in traceable amounts in the blood, Van Leeuwen surmised. To search for these blood-based biomarkers in a protein-rich serum, he developed a special selection approach. “We did not look in unbiased way. We made a first selection on the basis of the bone protein library (bone proteome) that we obtained from bone extracts, and linked that with a library of the serum proteins (serum proteome). This delivered a subset of 203 proteins, which is still quite a lot.” In the next selection this subset was linked to the gene expression profiles of bone-forming cells that they had gener-ated. This resulted in a short list of 50 candidate-biomarkers that are expressed by bone cells, present in the bone matrix and present in the circulation and which could predict an increased risk of fracture due to increased bone loss or poor bone quality.The NPC Valorisation Voucher makes it possible for the Van Leeuwen group to perform targeted MRM-MS protein assays to screen the final list of 50 candidate-biomarkers. “I expect that we will end up with a profile of biomarkers relating to osteoporosis and fractures. But if it turns out to be one specific biomarker, that is all right too.”

Milestones 2008 assessment of the plasma proteome 2008 assessment of the bone proteome2009 availability of well-documented extreme bone phenotype groups2009 gene expression profile of the bone-forming cell (osteoblasts)2010 NPC Valorisation Voucher2011... - analyse selection of 50 candidate-biomarkers - exploitation identified biomarkers by own company Arcarios BV or

sell knowledge to diagnostic industry

“Our findings provide a great market opportunity for the diagnostics of bone

diseases”Hans van Leeuwen

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Communications

Opening the window to the outside world

The NPC feels very strongly about communications, not only for the professional proteomics community, but also by familiarising society with proteomics and its applications in biotechnology and healthcare. Since March 200� these issues are addressed by NPC Communication Officer Martje Ebberink.

Martje Ebberink NPC Communication Officer: “Organising and developing communication activities comes down to teamwork”

“When I started my job, several communication activities had already been initiated,” Martje Ebberink says. She mentions the annual NPC

progress meeting, the NPC Highlights Magazine and the NPC website. “But these activities focus primarily on the professional proteomics community. It was my big challenge to develop and organise public events and education projects as well.” The NPC considers it necessary to establish educational activities and other means of communication to inform the public about proteomics and its crucial role in life sciences. Ebberink is enthusiastic about the many innovative applica-tions of proteomics in the biotechnical and biomedical field. “The societal impact is enormous,” she adds. To fulfil her task Ebberink started writing a thorough communication plan. “Distinguishing different targets for different groups and defining the goals are import aspects in such a plan.” The next step was translating the paper plans into practicable initia-

tives and achievable projects. Therefore Ebberink first sought contact with current activities at the NGI centres, especially the Centre for Society and Genomics (CSG). “In all its activi-ties, CSG seeks contact with society for interaction, dialogue and education. Insights gained from research are used to improve and renew public communication and education,” Ebberink explains.

Joining current initiativesMany of CSG’s communication and education projects are con-ducted in close cooperation with the genomics centres of the Netherlands Genomics Initiative (NGI), including the NPC. Ebberink was introduced to two projects already running: the website on genomics www.allesoverdna.nl and the ‘DNA-labs on the road’. “Both initiatives had already proven to be very successful. I contacted Frans van Dam, head of Communications at CSG, to discuss the contribution of the NPC.” It led to concrete actions to incorporate proteomics knowledge and technologies in both initiatives. In close collaboration with NPC researchers Reinout Raijmakers and Arjen Scholten (Utrecht University), Hienke Sminia (NBIC) and Nathalie Doorenweerd (NCMSB) Ebberink developed three lectures for the DNA-labs on the road, and she took a position on the editorial board of the website www.allesoverdna.nl. She emphasises the necessity for close cooperation with NPC researchers. “Based on my communica-tion skills I can design a concept, but the content has to be contributed by the researchers. It comes down to teamwork.”

Participating in science festivals“Besides using the lectures for the DNA-labs on the road, suc-cess of these is further garnered by using them in proteomics

“The enthusiastic response of the audience is really amazing”


About DNAThe website www.allesoverdna.nl provides an informative and inspiring environment where visitors can form their own opinion about genomics. The website is intended for everyone who is interested in genomics but focuses especially on upper-secondary students and their teachers. The website includes articles about more or less each and every aspect of genomics, but also ex-plains different sciences and technologies. A number of articles deal explicitly with the issue of proteomics. In the coming years the NPC wants to extend this.

October: Month of KnowledgeOktober Kennismaand (October Month of Knowledge) is the biggest science and technology event in the Netherlands. The event is organised annually from October 1 to October 31. A great variety of institu-tions, including the NPC, take part in this unique event to bring science and technol-ogy closer to the public. The 25th anniver-sary of the event was held in 2010. Almost 300,000 people visited the more than 150 venues with 900 different activities.

Initiatives for bringing proteomics to the people

Public communication & education

workshops,” says Ebberink. For example, these workshops are held at the National DNA lab day 2010 and during open university days. Today proteomics is also represented at public science festivals. Ebberink cites the renowned Dutch ‘Oktober Kennismaand’ (October Month of Knowledge). “Exciting dem-onstrations and eye-catching experiments during this month-long event allow people a glimpse into the fascinating world of science and technology. A different theme is highlighted every year.” NPC first participated in 2009. “It is amazing seeing the visitors — children as well as their parents — en-thusiastically taking part in this science festival.” NPC will again be present this year. Brainstorm sessions about attrac-tive experiments and workshops have already been started. “Finally, another great initiative is coming up next fall. It concerns a TV production, coordinated by NGI. In one of the

episodes, the NPC will introduce the viewer to the world of proteomics,” Ebberink says with pride.

Last but not least“We discussed many of our new initiatives in the field of public communication and education. But don’t forget our communi-cation activities for professionals,” Ebberink emphasises. “We do our best in each new issue of NPC Highlights to make the magazine worth reading and informative for the NPC network, including investigators and policymakers.” Much attention is also paid to the annual NPC progress meeting. Reputable (inter)national representatives are invited to present a keynote lecture. Furthermore, NPC informs the media by press releases. “Recently the press release about the EU grant for the PRIME-XS consortium resulted in several requests to talk about pro-teomics and our research projects,” Ebberink explains.

DNA labs on the roadA number of genomics centres have de-veloped ‘DNA-labs on the road’ offering practical lessons on genomics, intended for the higher classes of HAVO (higher general secondary education) and VWO (pre-univer-sity education). In these practical lessons, pupils are introduced to genomics topics, such as tumour profiling, bioinformatics and plant research, while a link is made to their own biological knowledge and contempo-rary genomics research. The DNA labs are a huge success and continue to develop new tools. In 2009 and 2010 the NPC developed three lectures on proteomics: Protein Next Top Model, Which protein are you? and Proteomics for the mass. www.dnalabs.nl

Lectures on proteomicsThe NPC lectures that have been developed for the DNA-labs on the road are also used separately for lectures at secondary schools. The lectures are given by researchers from the NPC, but are also suitable for presentation by the school teacher. Furthermore, workshops are organised during public university days.

National DNA lab dayThe National DNA lab day is aimed at teachers, technical research assistants and other interested parties. The participants are offered the opportunity to broaden their knowledge of the latest scientific developments, new didactics and the DNA-labs on the road. In addition to keynote speeches, there is a wide range of practical, didactic and substantive workshops available. In 2010 the NPC organised two workshops on proteomics.

TV seriesIn the fall of 2011 a series on genomics will be on Dutch television. An enterpris-ing presenter investigates the status quo in The Netherlands in the field of applying the latest DNA and proteomics technolo-gies. The broadcasts will show what we can expect in the near future based on current practices. In one of the episodes, the NPC will introduce the viewer to the world of proteomics.

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Matthias Mann (Max Planck Institute of Biochemistry, Martinsried, Germany) presented a keynote lecture during the NPC progress meeting 2011 (Utrecht, 13 January) entitled ‘High resolution, quantitative mass spectrometry: a toolbox for biomedical research’

Matthias Mann has an impressive curriculum vitae which includes numerous awards and prizes, and many of his publications have appeared in Nature,

Science or Cell. He got involved in mass spectrometry as a PhD student at Yale University, when he worked in John Fenn’s group. Mann contributed to the development of Electrospray Ionisation (ESI) mass spectrometry for which Fenn received the Nobel Prize in 2002. When asked about important milestones in his research career, Mann mentions his development of an initial algorithm for identifying mass spectrometrically fragmented peptides. After introduction of a mixture of peptide fragments into the mass spectrometer, a spectrum of peaks is generated, which are specific for the peptides in the mixture. “With this algorithm you only need the sequence of a partial peptide and its mass to compare it with a database in order to finally determine which protein is in the mixture. The algorithm has been im-proved since that time and today it makes comparison with a database a very fast procedure.”

SILAC technologyOne of the other important achievements by Mann and his research group is the development of the SILAC technology. This technology enables quantitative mass spectrometry. Since the absolute signal intensity of a peptide ion measured in an

MS run does not always reflect the abundance of the peptide present in the analysed sample, researchers have been looking for ways to more accurately measure the abundance of pep-tides. In the SILAC technology proteins are labelled by growing cells in media containing isotopically labelled amino acids. The labelled proteins serve as a control. By comparing the in-tensity of the signals of the labelled to the unlabelled proteins in the mixture, the relative quantity can be determined. Mann’s group also developed a SILAC mouse, which has become a very popular research tool. “One drawback of the SILAC technology is the fact that we cannot label humans,” says Mann. “But we have found a way to use SILAC to measure the relative abundance of proteins in human tissue. This is an important application since in the diagnosis of cancer the amount of expressed protein in a cancer facilitates the deci-sion for further treatment.”

Complete proteomeOne of the latest topics Mann has been working on is the de-velopment of a technique to analyse the complete proteome of organisms. This is difficult due to its enormous complexity and dynamic range. “With yeast we have already succeeded in determining almost the complete proteome consisting of more than 4,000 proteins. We have started with the human pro-teome, for which we have already analysed 10,000 proteins, but this still has to be developed further.”Mann feels it is very rewarding and challenging to develop new instrumentation for application in biological and medical science and to improve it each time. “As a biologist you pick a certain topic to study and you usually stick with it for many years. However, as a developer of instrumentation we meet a new application field almost every year. This way we get to know a lot of different research topics, which makes it very interesting.”

Mass spectrometry evolves to a vitally important proteomics tool

“My aim is to turn mass spectrometry into a user friendly, robust and reliable technology so that many people can use it,” says Professor Matthias Mann, director of the Department of Proteomics and Signal Transduction at the Max Planck Institute for Biochemistry. For the last 2� years Mann has been working on the development of instrumentation and analysis tools for many applications in the life sciences.

“We meet a new application field almost every year”


Lukas Käll is an assistant professor at the Centre for Biomembrane Research at the Stockholm University in Sweden. He spent time developing software for the

European lunar probe SMART-1 before deciding to take on a PhD in bioinformatics at the Karolinska Institute in Stockholm. Afterwards he moved to the University of Washington in Seattle (USA) for postdoctoral research with William Stafford Noble in the department of Genome Sciences. Upon his return to Sweden in 2008, Käll had the opportunity to start his own research group at the Stockholm University.

Inspired by a coffee machineDuring his time in Seattle, he developed a new algorithm for the post-processing of data obtained by shotgun proteomics experiments. “A traditional target database search cannot perfectly discriminate the good matches from the bad ones,” Käll explains. His new algorithm can improve the discrimina-tion between correct and incorrect peptide-spectrum matches from a collection of tandem mass spectra. Käll named the programme Percolator, in part because it reflects the iterative nature of the algorithm. “It circulates data just like water is circulated in a coffee machine.” But mostly, he says, because while he lived in the States his all-American neighbours in Seattle gave him a percolator to make what they considered decent coffee.

Percolator uses semi-supervised machine learning, in which a small amount of labelled data is used together with a large amount of unlabelled data to train the software. The matches from searching a decoy database provide the negative exam-ples for the classifier, and a subset of the high-scoring matches from the target database provide the positive examples. Käll summarises: “Percolator takes a set of different features from the peptide-spectrum matches and learns to optimally combine these features.” Percolator trains a machine learning algorithm called a support vector machine (SVM) to discrimi-nate between the correct and incorrect matches. The vector of features with their optimal weights is then used to re-rank matches from all queries, often leading to improved sensitiv-ity. Käll says: “At first I didn’t realise why Percolator gave such a performance boost. It roughly doubled the number of correct assignments, an improvement which is rarely seen in bioinformatics. We had spent quite some time trying to under-stand the imperfections of traditional scoring systems.”

ChallengesRecently, Käll has worked on the improvement of the Percolator algorithm by including the prediction of the retention time of the peptides in the spectra. By using sixty features of the peptides such as length, bulkiness and reten-tion time coefficient, the programme could predict 95 percent of the retention times correctly. Käll is proud of the result: “We improved 5-10 percent on a normal tryptic digest data set. In the best cases, we obtained a 25 percent performance increase on protease-K data sets. In addition, it can handle post-translational modification, which is a key issue in pro-teomics at the moment.” In the near future, Käll would like to expand his group and focus on the integration of proteomics data with other data such as transcriptomics. “I think both sides could benefit from the combination of their data. One of my goals is to start a cooperation between proteomics and transcriptomics research groups.”

New semi-supervised machine learning algorithm improves peptide identification

“The challenge is to understand the proteomics issues and to translate

them into bioinformatics problems”

“Bioinformatics bridges biology and computer science. It is a vibrant field that has recently come into focus,” explains Lukas Käll, a physicist who turned to bioinformatics. He develops new machine learning algorithms to improve the identification of peptides from mass spectra. Traditionally, peptide identification is done by a database search procedure in which the mass spectra of the peptides are matched. “These matches are scored to separate right matches from wrong matches, but these scores are often flawed.”

Lukas Käll (Centre for Biomembrane Research, Stockholm University,

Sweden) presented a keynote lecture during the NPC progress meeting

2011 (Utrecht, 13 January) entitled ‘Machine learning techniques to assess

confidence and increase yield for shotgun proteomics’

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Roland Annan (GlaxoSmithKline, Philadelphia, PA, USA) presented a keynote lecture during the NPC progress meeting 2011 (Utrecht, 13 January) entitled ‘Biological variability: a cardiac proteome study in the SILAC mouse’

Roland Annan stresses that the rapid development of mass spectrometric technologies in the last decade, applied to protein research, has catalysed entirely

new experimental approaches and addressed many new biological questions. “The question today is more like what not to work on?” When Annan started at GSK in the early nineteen nineties, research into cell signalling was rapidly developing. His research initially focussed on protein kinases and their substrates which played a role in phosphorylation-dependent signalling and cel-lular function. “We developed and improved MS techniques to study phosphorylation and how it affects signalling. Later, we expanded our research to other types of posttranslational modifi-cations of proteins, like methylation, acetylation and ubiquitina-tion, which have now all been shown to have important biological roles. Furthermore, we developed quantitative techniques to study protein function in signalling pathways.” Annan: “Part of this success is due to having a stable research group for the last twenty years. I am proud of the fact that our publication record is quite good. In a company, publication is often more difficult as it may conflict with intellectual property.”

Bad hypertrophyStudying the mouse cardiac proteome is one of the group’s quantitative proteome projects. Annan and his research group

investigate good and bad cardiac hypertrophy. More specifi-cally they focus on understanding heart enlargement, cardiac hypertrophy, through comparisons between models of health and disease. For this they compare the enlarged athletic heart (physiological hypertrophy) to heart enlargement associated with diseases (pathological hypertrophy). “Currently, the only comprehensive attempts are based upon mRNA studies,” says Annan. “However, since the presence of mRNA does not always predict whether the protein product is eventually produced, we focus on proteins.” The researchers hope to identify unique proteins involved in physiological hypertrophy as opposed to proteins involved in pathological hypertrophy. These proteins may represent new targets for drug development.

SILAC mouse“Since we want to know if certain proteins are expressed more under abnormal conditions in comparison to normal conditions, we need to compare signals of proteins in mass spectrometry.” Annan has chosen SILAC mice developed by Matthias Mann’s lab to help in these quantitative studies. In the SILAC mouse all lysine residues, in every protein, are labelled with stable (non-radioactive) heavy isotopes. “We use the cells of these mice as internal standards for all experi-ments.” In this way one can measure protein expression differ-ences under normal and abnormal conditions. The researchers compared exercising mice with sedentary mice and mice with arterial constriction (bands on their arteries) and looked at their proteomes. “Surprisingly, the number of protein changes between exercising mice and mice with pathological hyper-trophy is fairly small,” says Annan. “The situation is probably more complicated than we thought, but we will continue our study.”

Present proteomics technology opens up the field of biological questions

“The time for proteomics has come,” states Roland Annan, Associate Director of Molecular Discovery Research from GlaxoSmithKline. “After the human genome was sequenced, scientists discovered how much more complicated the human biological system is.” Proteomics will help unravel many of the biological questions. Mass spectrometry is a critical component in the analysis of proteins.

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Anne-Claude Gingras leads a signal transduction and proteomics lab at the Mount Sinai Hospital based Lunenfeld Research Institute in Toronto, Canada.

“We try to find out which protein binds to which protein. We do that on a small scale finding single protein’s binding part-ners, but also on a large scale studying an interaction network involving hundreds of proteins.” Studying these large interac-tomes enables more general conclusions about how proteins associate with one another. For example, by analysing not just phosphatases but the entire yeast kinase and phosphatase interactome, Gingras and co-workers found that kinases are much more likely to physically associate with other kinases than with anything else in the cell. “That helps us understand how signalling networks are organised. We would never have detected this by studying a single kinase.”

New algorithmThe proteins of interest are purified using affinity purifica-tion methods before analysis by mass spectrometry (AP-MS). This removes most of the background signals in the cell samples but not all. A data analysis programme is needed to subtract the remaining background. Gingras and colleague Alexey Nesvizhskii from the University of Michigan developed a method called Significance Analysis of INTeractome (SAINT). “We actually use the background to our advantage assuming

that it is reproducible. In that way we are able to use very crude purification approaches.” The algorithm relies on the fact that the peptides for each protein are sequenced more often in the mass spectrometer in the case of a purification of an interactor than in a control purification. Using the new algorithm, Gingras, Nesvizhskii and colleague Mike Tyers from the University of Edinburgh identified a kinase and phosphatase interaction network of almost 2,000 interac-tions in bakers’ yeast (Saccharomyces cerevisiae). The network was found to contain dense local regions of interactions that suggested new functions. In addition, Gingras explains, this type of physical interaction network complements parallel approaches to analyse genetic interactions: “Based on genetic interactions that the research group of Charlie Boone and colleagues in Toronto had found, it was predicted that many protein complexes and interactions had been missed by earlier analysis. We used our methods to search for these predicted orphan complexes. It has been great to be able to actually find them.”

CollaborationsGingras is happy with the collaborative research climate in Toronto, where resources are shared with the University of Toronto and a number of hospitals. A biochemist and mass spectrometrist by training, she is able to bridge the gap between these groups of researchers where communica-tion is sometimes hampered. “We purposely develop our new data analysis tools and methodology so that they are easy for others, mainly biologists, to use. We spend a lot of time developing visual interfaces and training people to use the programme.” More than a hundred researchers use the methods worldwide. “I am proud that we have been able to enable so many people to make their discoveries, even though sometimes I am not making the discoveries myself.”

Interactome studies reveal new phosphatase and kinase interaction networks

“We spend a lot of time developing visual interfaces and training

people to use the programme”

Phosphatases remove phosphate groups from proteins in contrast to kinases which add phosphate groups. Kinases are ten times more abundant and well known than phosphatases, but both play an equally important role in cell growth and proliferation, and thus in the development of diseases. “I would like to find out how phosphatases recognise their target. No one really knows. Finding out will be a huge achievement,” says Anne-Claude Gingras.

Anne-Claude Gingras (Samuel Lunenfeld Research Institute, Mount

Sinai Hospital, Toronto) presented a keynote lecture during the NPC progress meeting 2011 (Utrecht,

13 January) entitled ‘Tools for interaction proteomics: applications

to signalling networks’

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�0 | NPC Highlights Special | April 2011

European Collaborations

EU projects with contribution from NPC researchersPRIME-XSPRIME-XS is part of the Seventh Framework Programme (FP7) to support access to large-scale infrastructures in the Life Sciences. The goal of PRIME-XS is to provide access to state-of-the-art proteomics facilities to European scientists in the life sciences.Albert Heck, initiator of the consortium, brought together major European players in the field of proteomics. They provide access to their technology at six access facilities, distributed over Europe. In addition, the partners in the consortium de-velop new technologies to better aid the research community in answering current day scientific questions, and organise a wide range of meetings, courses and training events.

Duration: 2011-2015EU contribution: 8 M€Coordinator: Albert Heck, Utrecht University,

The NetherlandsNPC researchers: Albert Heck, Reinout Raijmakers, Shabaz

Mohammed, Utrecht UniversityPartners: twelve partners from nine countries:

The Netherlands, Belgium, Switzerland, Denmark, Spain, United Kingdom, Austria, France, Germany

BASIS - Breast Cancer Somatic Genetics StudyBreast cancer is the leading cause of cancer deaths in women and is the most common cause of all deaths in women aged under 40. Breast cancer is a heterogeneous dis-ease with a number of subtypes. Basis, a Seventh Framework Programme (FP7) project, will generate complete catalogues of somatic mutations in 500 breast cancers. These catalogues of mutations will give scientists a statistical power to iden-tify cancer genes that are mutated at a frequency of greater than 3 percent in this class of breast cancer. The results of this exhaustive and comprehensive set of studies will have a big impact on our understanding of the causes and biology of breast cancer.

Duration: 48 monthsEU contribution: 10.5 M€Coordinator: Mike Stratton, Wellcome Trust Sanger

Institute, UKNPC researcher: John Foekens, Erasmus MC RotterdamPartners: thirteen partners from eight countries:

UK, The Netherlands, USA, France, Norway, Sweden, Germany, Belgium

TORNADOTORNADO is a European Union funded large collaborative project under the Seventh Framework Programme (FP7).The TORNADO consortium combines a systemic and com-prehensive mechanistic approach with a focus on interdis-ciplinary skills provided by world leading experts working in close harmony with the food industry. TORNADO will enable broader applications, faster development, and more solid evidence for health claims based on detailed and thorough in depth assessment of underlying mechanisms relevant to health effects of dietary products compared to traditional approaches.

Duration: 2009-2013EU contribution: 5.6 M€Coordinator: Sven Patterson, Karolinska Institute,

SwedenNPC researcher: Hans Clevers, Hubrecht Institute, UtrechtPartners: twenty international partners

EuroSyStemEuroSyStem is an EU funded partnership between Universities, small to medium businesses (SMEs) and named researchers from both the public and private sector. The Centres involved are all international leaders in their respec-tive fields, and collectively provide the skills and technolo-gies necessary for ground-breaking stem cell research. There are 29 research groups involved in the project. The ambition of EuroSyStem is to interlink complementary biological and computational expertise so as to drive the generation of new knowledge on the characteristics of normal and abnormal stem cells. In addition, the EuroSyStem Project has created the European Stem Cell Group, a net-work of leading and emerging researchers who have become Associate Principal Investigators with the project. This group will continue to be a leading force in stem cell research collaboration and networking beyond the life of the current project.

Duration: 2008-2012EU contribution: 1 M€ Coordinator: Austin Smith, Cambridge University, UKNPC researcher: Hans Clevers, Hubrecht Institute, UtrechtPartners: twenty partners four based in the UK

and sixteen across seven other European countries.

INSTRUCTINSTRUCT is part of the European Strategy Forum on Research Infrastructures (ESFRI) and is funded by the Seventh Framework Programme (FP7). It will link the infor-mation obtained by the major structural biology methods with state-of-the-art cell biology techniques to provide a dynamic picture of key cellular processes at all scales with a focus on a few key biomedical questions and environment problems.Albert Heck (Utrecht University) and Carol Robinson (University of Oxford) are the coordinators of the INSTRUCT Associate Centre for Analysis of Protein Complexes by Mass Spectrometry.

In this NPC HighLights we provide a short list of

papers that appeared in some of the top journals

and to which NPC participants contributed

(2009 - 2010). With the guarantee of being by far

not comprehensive, this overview shows some

elegant ground-breaking research.

Orchestration of Floral Initiation by APETALA1

Kaufmann, K., Wellmer, F., Muiño, J.M., Ferrier, T., Wuest, S.E., Kumar, V., Serrano-Mislata, A., Madueño, F., Krajewski, P.,

Meyerowitz, E.M., Angenent, G.C., Riechmann, J.L. Science (2010) Apr 2;328(5974):85-9.

Business Unit Bioscience, Plant Research International, Wageningen 6700 AA, Netherlands.

PMID: 20360106

The MADS-domain transcription factor APETALA1 (AP1) is a key regulator of Arabidopsis flower development. To understand the molecular mechanisms underlying AP1 function, we identi-fied its target genes during floral initiation using a combina-tion of gene expression profiling and genome-wide binding studies. Many of its targets encode transcriptional regulators, including known floral repressors. The latter genes are down-regulated by AP1, suggesting that it initiates floral develop-ment by abrogating the inhibitory effects of these genes. Although AP1 acts predominantly as a transcriptional repressor during the earliest stages of flower development, at more advanced stages it also activates regulatory genes required for floral organ formation, indicating a dynamic mode of action. Our results further imply that AP1 orchestrates floral initiation by integrating growth, patterning, and hormonal pathways.

MONOPTEROS controls embryonic root initiation by regulating a mobile transcription factor

Schlereth, A., Moller, B., Liu, W., Kientz, M., Flipse, J., Rademacher, E.H., Schmid, M., Juergens, G., Weijers, D.

Nature (2010) Apr 8;464(7290):913-6Entwicklungsgenetik, Zentrum für Molekularbiologie der

Pflanzen (ZMBP), Universität Tübingen, Auf der Morgenstelle 3, 72076 Tübingen, Germany.

Acquisition of cell identity in plants relies strongly on positional information, hence cell-cell communication and inductive signalling are instrumental for developmental patterning. During Arabidopsis embryogenesis, an extra-em-bryonic cell is specified to become the founder cell of the primary root meristem, hypophysis, in response to signals from

adjacent embryonic cells. The auxin-dependent transcription factor MONOPTEROS (MP) drives hypophysis specification by promoting transport of the hormone auxin from the embryo to the hypophysis precursor. However, auxin accumula-tion is not sufficient for hypophysis specification, indicating that additional MP-dependent signals are required. Here we describe the microarray-based isolation of MP target genes that mediate signalling from embryo to hypophysis. Of three direct transcriptional target genes, TARGET OF MP 5 (TMO5) and TMO7 encode basic helix-loop-helix (bHLH) transcription factors that are expressed in the hypophysis-adjacent embryo cells, and are required and partially sufficient for MP-depen-dent root initiation. Importantly, the small TMO7 transcription factor moves from its site of synthesis in the embryo to the hypophysis precursor, thus representing a novel MP-dependent intercellular signal in embryonic root specification.

The immunoproteasome cleans up after inflammation

van Deventer, S., Neefjes, J.Cell (2010) Aug 20;142(4):517-8

The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands.

Immune cells and cells activated by the inflammatory cytokine interferon express variant proteasomes called immunopro-teasomes that are characterized by unique catalytic subunits. Seifert et al. (2010) now show in mouse models of inflam-matory disease that immunoproteasomes help prevent the accumulation of harmful protein aggregates.

Interrogating viral capsid assembly with ion mobility-mass spectrometry

Uetrecht, C., Barbu, I.M., Shoemaker, G.K., van Duijn, E.,

Heck, A.J.R.

Nature Chemistry (2011) Feb;3(2):126-32. Epub 2010 Dec 19

Biomolecular Mass Spectrometry and Proteomics Group,

Bijvoet Center for Biomolecular Research and Utrecht Institute

for Pharmaceutical Sciences, Utrecht University, Utrecht,

The Netherlands.

Most proteins fulfil their function as part of large protein complexes. Surprisingly, little is known about the pathways and regulation of protein assembly. Several viral coat proteins can spontaneously assemble into capsids in vitro with morphologies identical to the native virion and thus resemble ideal model systems for studying protein complex formation. Even for these systems, the mechanism for self-assembly is still poorly under-stood, although it is generally thought that smaller oligomeric structures form key intermediates. This assembly nucleus and larger viral assembly intermediates are typically low abundant

Top publications 2009 - 2010

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and difficult to monitor. Here, we characterised small oligo-mers of Hepatitis B virus (HBV) and norovirus under equilibrium conditions using native ion mobility mass spectrometry. This data in conjunction with computational modelling enabled us to elucidate structural features of these oligomers. Instead of more globular shapes, the intermediates exhibit sheet-like structures suggesting that they are assembly competent. We propose pathways for the formation of both capsids.

Ion mobility mass spectrometry of proteins and protein assemblies

Uetrecht, C., Rose, R.J., van Duijn, E., Lorenzen, K., Heck, A.J.R.Chem Soc Rev. (2010) May;39(5):1633-55

Biomolecular Mass Spectrometry and Proteomics Group,

Bijvoet Center for Biomolecular Research and Utrecht Institute

for Pharmaceutical Sciences, University of Utrecht, Padualaan

8, 3584 CH Utrecht, The Netherlands.

Traditionally, mass spectrometry has been a powerful analyti-cal method enabling the structural analysis of small mol-ecules, and later on peptides and proteins. With the advent of native mass spectrometry, using a combination of electrospray ionisation and time of flight analysis, mass spectrometry could also be applied to the mass determination of large protein complexes such as ribosomes and whole viruses. More recently, ion mobility has been coupled to mass spectrometry

providing a new dimension in the analysis of biomolecules, with ion mobility separating ions according to differences in size and shape. In the context of native mass spectrom-etry, ion mobility mass spectrometry opens up avenues for the detailed structural analysis of large and heterogeneous protein complexes, providing information on the stoichiom-etry, topology and cross section of these assemblies and their composite subunits. With these characteristics, ion mobility mass spectrometry offers a complementary tool in the context of structural biology. Here, we critically review the develop-ment, instrumentation, approaches and applications of ion mobility in combination with mass spectrometry, focusing on the analysis of larger proteins and protein assemblies.

An Oct4-centered protein interaction net-work in embryonic stem cells

van den Berg, D.L., Snoek, T., Mullin, N.P., Yates, A., Bezstarosti, K., Demmers, J., Chambers, I., Poot, R.A.

Cell Stem Cell (2010) Apr 2;6(4):369-81Department of Cell Biology, Erasmus MC,

Dr. Molewaterplein 50, Rotterdam, The Netherlands.

Transcription factors, such as Oct4, are critical for estab-lishing and maintaining pluripotent cell identity. Whereas the genomic locations of several pluripotency transcription factors have been reported, the spectrum of their interaction

partners is underexplored. Here, we use an improved affin-ity protocol to purify Oct4-interacting proteins from mouse embryonic stem cells (ESCs). Subsequent purification of Oct4 partners Sall4, Tcfcp2l1, Dax1, and Esrrb resulted in an Oct4 interactome of 166 proteins, including transcription factors and chromatin-modifying complexes with documented roles in self-renewal, but also many factors not previously associated with the ESC network. We find that Esrrb associated with the basal transcription machinery and also detect interactions be-tween transcription factors and components of the TGF-beta, Notch, and Wnt signaling pathways. Acute depletion of Oct4 reduced binding of Tcfcp2l1, Dax1, and Esrrb to several target genes. In conclusion, our purification protocol allowed us to bring greater definition to the circuitry controlling pluripotent cell identity.

Phosphorylation dynamics during early differentiation of human embryonic stem cells

Van Hoof, D., Muñoz, J., Braam, S.R., Pinkse, M.W., Linding, R., Heck, A.J.R., Mummery, C.L., Krijgsveld, J.

Cell Stem Cell (2009) Aug 7;5(2):214-26Developmental Biology and Stem Cell Research, Hubrecht

Institute, Utrecht, The Netherlands.

Pluripotent stem cells self-renew indefinitely and possess characteristic protein-protein networks that remodel dur-ing differentiation. How this occurs is poorly understood. Using quantitative mass spectrometry, we analyzed the (phospho)proteome of human embryonic stem cells (hESCs) during differentiation induced by bone morphogenetic protein (BMP) and removal of hESC growth factors. Of 5222 proteins identified, 1399 were phosphorylated on 3067 residues. Approximately 50% of these phosphosites were regulated within 1 hr of differentiation induction, revealing a complex interplay of phosphorylation networks spanning different signaling pathways and kinase activities. Among the phos-phorylated proteins was the pluripotency-associated protein SOX2, which was SUMOylated as a result of phosphorylation. Using the data to predict kinase-substrate relationships, we reconstructed the hESC kinome; CDK1/2 emerged as central in controlling self-renewal and lineage specification. The find-ings provide new insights into how hESCs exit the pluripotent state and present the hESC (phospho)proteome resource as a complement to existing pluripotency network databases.

Other highlighted publications

Chughtai, K., Heeren, R.M.A.Mass Spectrometric Imaging for biomedical tissue analysisChem Rev. (2010) May 12;110(5):3237-77 (review)

Muñoz, J., Heck, A.J.R. Snapshots of kinase activitiesNat Biotechnol. (2009) Oct;27(10):912-3

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�2 | NPC Highlights Special | April 2011

Jansen, M., Ten Klooster, J.P., Offerhaus, G.J., Clevers, H.H. LKB1 and AMPK Family Signaling: The Intimate Link Between Cell Polarity and Energy MetabolismPhysiol Rev. (2009) 89: 777-798

Landeira, D., Sauer, S., Poot, R., Dvorkina, M., Mazzarella, L., Jørgensen, H.F., Pereira, C.F., Leleu, M., Piccolo, F.M., Spivakov, M., Brookes, E., Pombo, A., Fisher, C., Skarnes, W.C., Snoek, T., Bezstarosti, K., Demmers, J., Klose, R.J., Casanova, M., Tavares, L., Brockdorff, N., Merkenschlager, M., Fisher, A.G.Jarid2 is a PRC2 component in embryonic stem cells required for multi-lineage differentiation and recruitment of PRC1 and RNA Polymerase II to developmental regulatorsNature Cell Biology (2010) Jun;12(6):618-24

Van den Bogaart, G., Meinema, A.C., Krasnikov, V., Veenhoff, L.M., Poolman, B. Nuclear Transport factor directs localization of protein synthesis during mitosisNature Cell Biology (2009) Mar;11(3):350-6

Witte, M. D., Kallemeijn, W. W., Aten, J., Li, K.-Y., Strijland, A., Donker-Koopman, W.E., Blijlevens, B., Kramer, G., van den Nieuwendijk, A. M. C. H., Florea, B.I., Hooibrink, B., Hollak, C. E. M., Ottenhoff, R., Boot, R. G., van der Marel, G.A., Overkleeft, H. S., Aerts, J. M. F. G. Ultrasensitive in situ visualization of active glucocerebrosidase moleculesNat Chem Biol. (2010) Dec;6(12):907-13

Geiger, S.R., Lorenzen, K., Schreieck, A., Hanecker, P., Kostrewa, D., Heck, A.J.R., Cramer, P.RNA polymerase I contains a TFIIF-related DNA-binding subcomplexMol. Cell (2010) Aug 27;39(4):583-94

Moshkin, Y.M., Kan, T.W., Goodfellow, H., Bezstarosti, K., Maeda, R.K., Pilyugin, M., Karch, F., Bray,S.J., Demmers, J.A., Verrijzer, C.P. Histone chaperones ASF1 and NAP1 differentially modulate removal of active histone marks by LID-RPD3 complexes during NOTCH silencing Mol. Cell (2009) Sep 24;35(6):782-93PMID: 19782028

ten Klooster, J.P., Jansen, M., Yuan, J., Oorschot, V., Begthel, H., Di Giacomo, V., Colland, F., de Koning, J., Maurice, M.M., Hornbeck, P., Clevers, H. Mst4 and Ezrin induce brush borders downstream of the Lkb1/Strad/Mo25 polarization complexDev Cell. (2009) 16: 551-562

Jaworski, J., Kapitein, L.C., Gouveia, S.M., Dortland, B.R., Wulf, P.S., Grigoriev, I., Camera, P., Spangler, S.A., Di Stefano, P., Demmers, J., Krugers, H., Defilippi, P., Akhmanova, A., Hoogenraad, C.C.Dynamic microtubules regulate dendritic spine morphology and synaptic plasticityNeuron (2009) Jan 15;61(1):85-100

Mahmoudi, T., Boj, S.F., Hatzis, P., Li, V.S., Taouatas, N., Vries, R.G., Teunissen, H., Begthel, H., Korving, J., Mohammed, S., Heck, A.J.R., Clevers, H.The leukemia-associated Mllt10/Af10-Dot1l are Tcf4/β-catenin coactivators essential for intestinal homeostasisPLoS Biology (2010) Nov 16;8(11):e1000539

Kaufmann, K., Muiño, J.M., Jauregui, R., Airoldi, C.A., Smaczniak, C., Krajewski, P., Angenent, G.C.Target genes of the MADS transcription factor SEPALLATA3: Integration of developmental and hormonal pathways in the Arabidopsis flowerPloS Biology (2009) Apr 21;7(4):e1000090

Jore, M. M., Lundgren, M., van Duijn, E., Bultema, J.B., Westra, E.R., Waghmare, S.P., Wiedenheft, B., Pul, Ü., Wurm, R., Wagner, R., Beijer, M.R., Barendregt, A., Zhou, K., Snijders, A.P., Dickman, M.J., Doudna, J.A., Boekema, E.J., Heck, A.J.R., van der Oost J., Brouns, S.J.Structural basis for CRISPR RNA-guided DNA recognition by CascadeNat Struct Mol Biol. (2010).

Spitzer, J., Poolman, B. The Role of Biomacromolecular Crowding, Ionic Strength and Physicochemical Gradients in the Complexities of Life’s EmergenceMicrobiol. Molec. Biol. Rev.(2009) 73, 371-388

Brettschneider, C., Rose, R.J., Hertel, S., Axmann, I. M., Heck, A.J.R., Kollmann, M.A sequestration feedback determines dynamics and temperature entrainment of the KaiABC circadian clockMol Syst Biol. (2010) Jul 13;6:389

van Wijk, S.J., de Vries, S.J., Kemmeren, P., Huang, A., Boelens, R., Bonvin, A.M., Timmers, H.Th.M.A comprehensive framework of E2-RING E3 interactions of the human ubiquitin-proteasome systemMol Syst Biol. (2009) 5:295

Geurink, P.P., Florea, B.I., Li, N., Witte, M.D., Verasdonck, J., Kuo, C.L., van der Marel, G.A., Overkleeft, H.S.A cleavable linker based on the levulinoyl ester for activity-based protein profiling Angew. Chem. Int. Ed. (2010) Sep 10;49(38):6802-5

Hillaert, U., Verdoes, M., Florea, B.I., Saragliadis, A., Habets, K.L., Kuiper, J., Van Calenbergh, S., Ossendorp, F., van der Marel, G.A., Driessen, C., Overkleeft, H.S.Receptor-mediated targeting of cathepsins in professional antigen presenting cellsAngew. Chem. Int. Ed. (2009) 48, 1629

Tora, L., Timmers, H.Th.M.The TATA box regulates TBP dynamics in vivoTIBS. (2010) 35:309-314.

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Quantitative proteomics on the fly

Joost GouwUtrecht University, 14 January 2009Promotor: Prof. dr. A.J.R. HeckCo-promotor: Dr. J. Krijgsveld

Gene expression Profiling of Early Stage Non-Small Cell Lung Cancer

Jun HouErasmus MC, 13 January 2010Promotores: Prof. dr. J.N.J.Philipsen and Prof. dr. F.G.Grosveld

Genome-wide regulation of TATA-binding activity

Folkert van WervenUMC Utrecht, 20 January 2009Promotor: Prof. dr. H.Th.M. Timmers

Biomarker Discovery for head and neck cancer. A proteomics approachTieneke Schaaij – VisserUtrecht University, 20 January 2010Promotores: Prof. dr. A.J.R. Heck and Prof. dr. R.H. BrakenhofCo-promotor: Dr. M. Slijper

Chromatin structure in cell differentiation, aging and cancer

Sima Kheradmand KiaErasmus MC, 3 June 2009Promotor: Prof. dr. C.P. Verrijzer

Probing the drug interactome by chemical proteomics

Poupak DadvarUtrecht University, 29 October 2009Promotor: Prof. dr. A.J.R. Heck

Functional and mechanisitic aspects of endogenous RNAi in C. elegansJosien van WolfswinkelUtrecht University, 16 December 2009Promotores: Prof. dr. A.J.R. Heck and Prof. dr. ir. E.P.J.G. CuppenCo-promotores: Dr. Jeroen Krijgsveld, Dr. R.F. Ketting

Activity-based profiling of glycoconjugate processing enzymesMartin WitteLeiden University, 22 December 2009Promotores: Prof. dr. H.S. Overkleeft and Prof. dr. G.A. van der MarelCo-promotor: Dr. B.I. Florea

Redox proteomics sheds light on photodynamic treatment of cancer

Pavel TsaytlerUtrecht University, 28 January 2009Promotor: Prof. dr. M.R. Egmond

200�

2010

Nuclear Receptor and Multiple Endocrine Neoplasia Type 1 MEN1

Koen DreijerinkUMC Utrecht, 25 June 2009Promotores: Prof. dr. H.Th.M. Timmers and Prof. dr. C.J.M. Lips

Advancing Liquid Chromatography-Mass Spectrometry based technologies for proteome researchPaul BoersemaUtrecht University, 27 January 2010Promotor: Prof. dr. A.J.R. HeckCo-promotor: Dr. S. Mohammed

Selectivity of E2E� interactions in the human ubiquitin system

Sjoerd van WijkUMC Utrecht, 28 January 2010Promotor: Prof. dr. H.Th.M. Timmers

PhD Theses

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Structure and function of substrate-binding domains of ABC transporters

Ronnie BerntssonUniversity of Groningen, 23 April 2010Promotores: Prof. dr. B. Poolman and Prof. dr. D.J. Stolboom

Quantitative proteomics of Saccharomyces cerevisiae vacuoles and stress responses in Lactococcus lactis Elena WiederholdUniversity of Groningen, 10 May 2010Promotores: Prof. dr. D.J. Slotboom and Prof. dr. B. Poolman

MADS on the move

Susan UrbanusWageningen University, 19 May 2010Promotor: Prof. dr. ir. G.C. AngenentCo-promotor: Dr. ir. R.G.H. Immink

Chemical Phosphoproteomics and Development of Bisubstrate Based Inhibitors of Protein Kinase C IsozymesAlex PootUtrecht University, 19 May 2010Promotor: Prof. dr. Rob LiskampCo-promotores: Dr. ir. D.T.S. Rijkers and Dr. M. Slijper

The antifungal activity of Natamycin. A novel model of action of the polyene antibioticsYvonne te WelscherUtrecht University, 16 June 2010Promotor: Prof. dr. B. de KruijffCo-promotor: Dr. E.J. Breukink

Carbohydrate-based chemical probes for the proteomic profiling of glucosidases and the emerging cancer marker galectin-�

Monique van ScherpenzeelUtrecht University, 15 February 2010Promotor: Prof. dr. R.M.J. Liskamp

RPTPa-mediated activation of Src

Andrei VacaruHubrecht Institute, Utrecht, 18 February 2010Promotor: Prof. dr. H. Clevers

A proteomics approach to inner membrane biogenesis in Escherichia coli

Claire PriceUniversity of Groningen, 26 February 2010Promotor: Prof. dr. A.J.M. Driessen

Monolithic Liquid-Chromatography c=Columns for Protein Analysis

Michiel van de MeentUtrecht University, 16 March 2010Promotor: Prof. dr. G.J. de Jong

Phosphorylation and Ubiquitination in Growth Hormone Receptor Endocytosis and Signalling

Joyce PuttersUtrecht University, 16 April 2010Promotor: Prof. dr. G.J.A.M. Strous

Divide and conquer strategies for the in-depth analysis of proteomes

Sharon GauciUtrecht University, 8 September 2010Promotor: Prof. dr. A.J.R. HeckCo-promotor: Dr. J. Krijgsveld

2010

Probing yeast molecular systems biology by new proteomics strategies

Andreas HelbigUtrecht University, 13 September 2010Promotor: Prof. dr. Albert HeckCo-promotor: Dr. Monique Slijper

Application of Chemical Proteomics to Biomarker Discovery in Cardiac Research

Thin Thin AyeUtrecht University, 15 September 2010Promotor: Prof. dr. A.J.R. HeckCo-promotor: Dr. A. Scholten

Writers and readers of histone H� methylation

Radhika WarrierUMC Utrecht, 23 September 2010Promotor: Prof. dr. H.Th.M. Timmers

PhD Theses

| ��

On the mechanism of prokaryotic glutamate transporter homologues

Maarten GroeneveldUniversity of Groningen, 1 October 2010Promotor: Prof. dr. D.J. Slotboom

Probing proteasome activity and functionCancer diagnostics and mechanism of antigen processing

Celia Berkers (cum laude)NKI, Amsterdam, 5 October 2010Promotores: Prof. dr. H.S. Overkleeft and Prof. dr. J. NeefjesCo-promotor: Dr. H. Ovaa

Synthetic tools to illuminate matrix metalloproteinase and proteasome activitiesPaul Geurink (cum laude)Leiden University, 6 October 2010Promotores: Prof. dr. H.S. Overkleeft and Prof. dr. G.A. van der MarelCo-promotor: Dr. B.I. Florea

Beyond bottlenecks in membrane protein production

Ravi K.R. MarreddyUniversity of Groningen, 15 October 2010Promotor: Prof. dr. B. Poolman

Profiling of ABC transporters

Karin WoltersUniversity of Groningen, 15 October 2010Promotor: Prof. dr. B. Poolman

Lateral diffusion of proteins and peptides in membranes

Siva RamaduraiUniversity of Groningen, 18 October 2010Promotor: Prof. dr. B. Poolman

Function and regulation of the histone methyltransferase Dot1

Floor FrederiksNKI Amsterdam, 29 October 2010Promotor: Prof. dr. M.M.S van Lohuizen Co-promotor: Dr. F. van Leeuwen

Analysing macromolecular structures with native mass spectrometry: insights to virus structure and assembly

Charlotte Uetrecht (cum laude)Utrecht University, 24 November 2010Promotor: Prof. dr. A.J.R. Heck

Composition and function of Ccr�-Not protein complexes

Nga-Chi LauUMC Utrecht, 30 November 2010Promotor: Prof. dr. H.Th.M. Timmers

Lys-N: a versatile enzyme for proteomics

Nadia TaouatasUtrecht University, 20 December 2010Promotor: Prof. dr. A.J.R. HeckCo-promotor: Dr. S. Mohammed

Unravelling the molecular mechanisms of the canonical Wnt signaling pathway

Ser Sue NgHubrecht Institute Utrecht, 28 September 2010Promotor: Prof. dr. H. CleversCo-promotor: Dr. T. Mahmoudi

2010

PhD Theses

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Groningen

Amsterdam

Utrecht Wageningen

RotterdamNijmegen

Leiden

Delft

High level proteomics research, technology development & transfer for biological and biomedical applications

The Netherlands Proteomics Centre (NPC) is a strategic collaboration of research groups from universities, academic medical centres and (biotech) companies. With a scientific programme addressing key areas of proteomics in several projects and specialised Research Hotels, the NPC performs high-quality research and knowledge transfer in an international context.

Netherlands Proteomics Centre At a glance

Core Consortium Participants

• Utrecht University (UU)

• University Medical Centre Utrecht (UMCU)

• University of Groningen (RUG)

• Leiden University (LU)

• Leiden University Medical Centre (LUMC)

• Delft University of Technology (TU Delft)

• Radboud University Nijmegen (RUN)

• University of Amsterdam (UvA)

• Academic Medical Centre Amsterdam (AMC)

• Wageningen University and Research Centre/

Plant Research International (WUR/PRI)

• Erasmus Medical Centre Rotterdam (EMC)

• Netherlands Cancer Institute (NKI-AVL)

• National Institute for Public Health and the

Environment (RIVM)

• FOM Institute AMOLF

• Hubrecht Institute

Affiliated Companies

• Agilent Technologies• Arcarios• Artemis Pharmaceuticals• BIOBASE GmbH• Bioceros• Bruker Daltonics B.V• Bruker Nonius• Carls Zeiss• Cellzome AG• CiberNed• DSM• deCODE Genetics• Eli Lilly• Epigeomics AG• FEI Electron Optics• Galapagos• Genmab• Harbour Antibodies• Hybrigenics• IBM• Jansen Pharma• Johnson and Johnson• Merus B.V.• Novartis• Organon/Merck• PamGene B.V.• Pfizer• Philips• Pluriomics• Profibrix• Roche• Source BioScience• Thermo Scientific• UbiQ• Veridex LLC

Netherlands Genomics Initiative

NGI creates societal and economic value from ge-nomics through a network of large-scale Genom-ics Centres that work on targeted programmes. Excellent research, state-of-the-art technology infrastructure, societal embedding of genomics and a stimulating environment for research talent are key elements of the NGI strategy.

NGI Genomics Centres

Sixteen NGI Genomics Centres are responsible for breakthroughs in the fields of health, agro-food, sustainability, enabling technologies, safety and society. The Genomics Centres are consortia of universities, research institutes, industry and societal organisations. The NPC is part of the enabling technologies cluster, consisting of the following centres:

netherlands centre

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NPC T1 Cancer Proteomics – G.J.P.L. Kops (UMCU)NPC T2 Proteome Biology of Plants – G.C. Angenent (PRI), B. Scheres (UU)NPC T� Proteome Biology of Micro-organisms – P.D.E.M. Verhaert (TUD), B. Poolman (RUG)NPC T� Proteome Biology of Stem cells – H.Th.M. Timmers (UMCU), A.J.R. Heck (UU)NPC T� Proteomics and Autoimmune Diseases – F. Koning (LUMC)

NPC E1 New Separation and Enrichment Tools in Proteomics – S. Mohammed (UU), R.P.H. Bischoff (RUG)

NPC E2 Chemical Approaches to Proteome Biology – H.S. Overkleeft (LU), H. Ovaa (NKI-AVL)

NPC E� New Mass Spectrometric Tools in Proteomics – R.M.A. Heeren (FOM-AMOLF), A.J.R. Heck (UU)NPC E� Bioinformatics in Proteomics – B. van Breukelen (UU), P. Horvatovich (RUG)

Analytical Hotel Utrecht – A.J.R. Heck (UU), A.F.M. Altelaar (UU)Analytical Hotel Groningen – B. Poolman (RUG), F. Fusetti (RUG)Analytical Hotel Delft – P.D.E.M. Verhaert (TUD), M.W. Pinkse (TUD)Analytical Hotel Rotterdam – J. Demmers (EMC)Plant Proteomics Wageningen – G.C. Angenent (PRI), A.H.P. America (PRI)Bioinformatics Hotel Utrecht – B. van Breukelen (UU)Cell Sorting Hotel Utrecht – B. Scheres (UU)Cell Culture Hotel Utrecht – H.Th.M. Timmers (UMCU), P.W. Pijnappel (UMCU)

The NPC has started, together with the Netherlands Bioinformatics Centre (NBIC), several bioinformatics projects to produce generic tools for proteomics data storage and analysis.NPC-GM T1 Information management and processing

infrastructure - M. Swertz (UMCG), A. Stubbs (EMC)NPC-GM T2 Data processing: from raw data to consistently

annotated peak-intensity tables - A.H.P America (PRI), B. Van Breukelen (UU)NPC-GM T3 Functional annotation and classification

- A. Stubbs (EMC), A.H.C. van Kampen (UvA)

The NPC promotes knowledge transfer and valorisation. In close collaboration with local Tech Transfer Offices the NPC offers coaching and the NPC Valorisation Voucher, a Proof-of-Concept Fund to bring technologies to the market.

Valorisation

NPC Research HotelsNPC Research Themes

NPC/NBIC Gaining Momentum

NPC Enabling Technologies

Mission

It is the mission of the Netherlands Proteomics Centre (NPC) to shape coherence and collaboration in proteomics research in the Netherlands. Thereby, the NPC wants to continue to be recognised as an internationally outstanding partner in proteomics research in Europe and beyond.The NPC will improve and widen fundamental technological tools in proteomics and proteomics related bioinformatics research, and through well-organised knowledge transfer will make them available to the biological and biomedical research communities at large. With the help of these new tools proteomics research will contribute to a better understanding of processes of health and disease, and pave the way for the generation of new means for intervention and prevention of undesired life-processes.


2

39 3

2

16

6

3

7

2

2

33

9

17

International collaborationsNumber*

Europe 132

China 1

Hawaii 1

Japan 3

Mexico 1

Nigeria 1

Taiwan 2

USA 46

Collaborations with NGI Centres of ExcellenceNumber of projects

Netherlands Bioninformatics Centre 21

Centre for Society & Genomics 4

Cancer Genomics Centre 42

Celiac Disease Consortium 4

Centre for Biosystems Genomics 31

Centre for Medical Systems Biology 2

Kluyver Centre for Genomics of Industrial Fermentation 31

VIRGO 6

Netherlands Consortium for Systems Biology 3

1

2

2

34

56

108

52

135

National collaborations

Number*: 390

* As evidenced by co-authorship of NPC scientific publications

NPC Scientific Network

The NPC forms a national platform for the development and application of proteomics technolo-gies that is extensively linked with national and international research groups in the life sciences. This page provides an overview of collaborations with other NGI Centres of Excellence, and with other national and international research groups. These collaborations are established via the NPC scientific programme as well as through the NPC Research Hotels.


Facts and Figures 2009 - 2010Netherlands Proteomics Centre

IF > 105 < IF < 10IF < 50

20

40

60

80

Num

ber

0

50

100

150

200

PhD thesesOral presentationsPostersArticles

Num

ber

0

1

2

3

4

NPC Vouchersgranted

Spin-off'sPatents

Num

ber

0

5

10

15

20

25

30

35

Educationalactivities

Oralpresentations

Publications

Num

ber

0

4

8

12

16

With industryWith academiaand hospitals

Valu

e (M

�)

2009

2010

Impact factor scientific articles

Newly acquired projects

Valorisation output

Non-scientific output

Scientific output

0

40

80

120

160

Training projectsResearch projects

Num

ber

Hotel trainings and projects

Facts and Figures 2009 - 2010Netherlands Proteomics Centre

Full professors (2%)Associate professors/assistant professors (5%)

Postdocs(35%) PhD students

(44%)

Technicalstaff(14%)

Management & communication (4%)

Equipment(21%)

Scientific personnel(75%)

Research funds(NWO, KNAW, ESF)

(11%)

Privatefunding(11%)

Direct fundingby institutes

(35%)

NGI subsidy(43%)

2009

2010

NPC Funding

NPC Expenditures

NPC Research Personnel

NPC Achievements

• State of the art infrastructure

• National coordination via research themes

• Development of innovative proteomics approaches

• High level scientific output

• Intensive knowledge transfer via Research Hotels

• Business coaching and proof-of-concept funding

• Strong industrial and academic liaisons

• Integrated bioinformatics programme

• Increased awareness for public and societal aspects

• Effectively embedded in National Genomics Initiative

• World-wide scientific network

• Links with international proteomics organisations

Total 21,648 Ke

Total 21,648 Ke

Total 144,7 fte

Over the last years, many opinion leaders from outside the NPC have shared their views on proteomics and the NPC in particular at this place. Please read a view memorable quotes below.

“The NPC is one of our technology-developing powerhouses within the Netherlands Genomics Initiative (NGI). Increasingly NPC, and the other NGI centres, are recognised in the international R&D community as centres of excellence, where public and private partners collaborate to translate science into products of economic and societal value.” Dr. Colja Laane, Director Netherlands Genomics Initiative

59th ASMS Conference on Mass Spectrometry, Denver, USA

Bessensap, Museon, The Hague, The Netherlands

5th European Summer School “Proteomic Basics”, Brixen/Bressanone, South Tirol, Italy

HUPO 2011, Geneva, Switzerland

Life Sciences Momentum 2011, The Netherlands

upcoming events5-9 June 2011

6 June 2011

31 July - 6 August 2011

4-7 September 2011

22 November 2011

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“Progress in systems genetics depends critically on a close interaction between biologists and technology development centres such as NPC. We wish that the cooperation with NPC will continue in the future.” Dr. Ernst Hafen, Institute of Molecular Systems Biology, ETH Zürich and CEO evalueSCIENCE

“Clearly proteomics remain a highly dynamic area of research, but the unique NPC concept, which brings together high quality proteomics research facilities, will be the best guarantee for remaining at the leading edge of science.” Dr. Jan van Oostrum, Novartis

“The NPC, founded on scientific excellence and well-positioned for knowledge transfer to industrial partners, plays an important role in the vital area of Molecular Medicine, paving the way to the future of healthcare”. Dr. Hans Hofstraat, vice president research, Philips

“The intense collaboration with the bioinformatics team at the NPC will certainly lead to a fruitful cross-fertilisation, and can be seen as a model for future international partnerships at the interface of proteomics and bioinformatics”. Dr. Rolf Apweiler, European Bioinformatics Institute (EBI), president Human Proteome Organisation (HUPO)

“The flexible collaborative concept of the NPC leads to scientific excellence, development of new technologies and sustainable resources such as facilities. The NPC had indeed achieved an impact beyond its original goals and reached an international level of excellence.” Dr. Hans-Werner Mewes, Chair review committee NPC phase I, Max-Planck-Institut Martinsried

“Proteomics can become a valuable tool to catch the unlimited diversity of molecules in nature, areas which could have tremendous value for the benefit of mankind. The NPC, might be very well positioned to play a leading international role in this endeavor.” Dr. Joël Vanderkerckhove, Flanders Institute for Biotechnology (VIB)

“As proteomics research is inherently complex in nature and rapidly evolving, the NPC concept of the Research Hotels is brilliant, as it allows novice researchers to learn from experts over an extended period of time.” Dr. Cindy Bell, vice president Genome Canada

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npc highlights 13 special | April 2011 PECIAL · Palma (UU). The PhD students can use their prize...

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