1406 VOLUME 21 | NUMBER 12 | DECEMBER 2015 nature medicine
p e r s p e c t i v e F o c u s o n AG I n G
Accumulation of intracellular damage is an almost universal hallmark of aging. An improved understanding of the systems that contribute to cellular protein quality control has shed light on the reasons for the increased vulnerability of the proteome to stress in aging cells. Maintenance of protein homeostasis, or proteostasis, is attained through precisely coordinated systems that rapidly correct unwanted proteomic changes. Here we focus on recent developments that highlight the multidimensional nature of the proteostasis networks, which allow for coordinated protein homeostasis intracellularly, in between cells and even across organs, as well as on how they affect common age-associated diseases when they malfunction in aging.
Protein homeostasis, or proteostasis, is assured through the coordi-nated action of intricate cellular systems—the proteostasis networks. Under normal conditions, these systems rapidly sense and rectify disturbances in the proteome to restore basal homeostasis1. During stress, similar systems preserve proteome solubility and functionality by bringing it to an altered point of proteostasis balance that takes into consideration the stress-induced cellular changes2.
Although the robustness and adaptability of the proteostasis net-works is remarkable, if stressors are chronic, the proteostasis balance becomes difficult to maintain, and proteotoxicity develops3. With age, the ability of many cells and organs to preserve proteostasis under resting and stress conditions is gradually compromised4. Loss of proteostasis is part of the pathogenesis of many human patholo-gies, including neurodegenerative diseases such as Alzheimer’s disease or Parkinson’s disease3. It is not a coincidence that many of these diseases—generically known as proteinopathies or protein confor-mational diseases—are regarded as age-related disorders, given that the physiological deterioration of the proteostasis networks with age is an important aggravating factor in these diseases1,4,5.
Numerous lines of evidence support a tight relationship between proteostasis and healthy aging. Although a gradual loss of proteos-tasis can be detected in most organisms as they age, the longest-living species have been shown to have more stable proteomes6 (comprised of cellular proteins that are more resistant to damage), and, for example, in the case of the long-lived naked mole rat, pro-teome stability correlates with enhanced activity in the proteostasis systems7. Furthermore, interventions that modulate the activity
of the proteostasis networks in invertebrates and in mammals extend their lifespans and healthspans1,3,4.
A detailed description of each of the systems that maintains cellular proteostasis is beyond the scope of this Perspective. Here we focus on the recognized importance of the loss of proteostasis in aging. We first describe specific characteristics of the proteostasis networks that make these systems vulnerable to the chronic stress that is often associated with aging. Then we touch upon newly identified dimen-sions of the proteostasis networks, beyond the relatively well-known cellular cytoplasm, that have transformed the ways in which we think about proteostasis. This includes concepts such as organelle proteostasis, organ or tissue proteostasis and even organism pro-teostasis networks that help to integrate a coordinated proteostatic response throughout the whole body. We argue that this integrated view of proteostasis is of utmost relevance to further understand the basis and consequences of a loss of proteostasis in aging and that it has generated considerable interest as a yet-unexplored therapeutic target for the treatment of age-related diseases.
Aging of the components of proteostasis systemsThe main players in proteostasis maintenance are chaperones and two proteolytic systems, the ubiquitin-proteasome and the lysosome- autophagy systems (Fig. 1). These components decide the fate of unfolded proteins: whether they will refold into their original stable conformation or whether they will instead be eliminated from the cell through proteolysis8.
Chaperones. Chaperones assist proteins through each of the different conformational changes that they undergo during their lifetime, which include de novo folding, assembly and disassembly, transport across membranes and targeting for degradation9. The need for chaperones originates from the crowded environment in the cytoplasm as well as in the lumen of most organelles. An important aspect of chaperone functioning is the molecules’ ability to integrate multiple cellular cues in order to decide the fate of cellular proteins that have lost their stable conformations. Thus, even for proteins that have experienced the same degree of unfolding, chaperones will assist them to either refold or degrade, depending on the feasibility of the cell doing one or the other at that particular time. Factors such as cellular ATP content (because substrate binding and release requires ATP hydrolysis), as well as over-all chaperone availability, may also contribute to this final decision.
Once a commitment to degradation is made, chaperones often also decide the proteolytic pathway that each misfolded protein will follow. For example, HSC70, a constitutive cellular chaperone, can target pro-teins for degradation. The two systems most commonly used for this breakdown of proteins are the proteasome and autophagy systems. The proteasome, a multi-subunit protease that is most abundant in
Department of Developmental and Molecular Biology, Institute for Aging Studies, Albert Einstein College of Medicine, New York, New York, USA. Correspondence should be addressed to A.M.C. ([email protected])
Received 20 September; accepted 2 November; published online 8 December 2015; doi:10.1038/nm.4001
Proteostasis and agingSusmita Kaushik & Ana Maria Cuervo
nature medicine VOLUME 21 | NUMBER 12 | DECEMBER 2015 1407
the cytosol—although it can also be detected in the nucleus—is responsible for the rapid degradation of proteins that are often tagged with covalently attached strands of the small protein ubiquitin10,11. Proteins can also be degraded in lysosomes through a process known as autophagy. Different types of autophagy have been identified (macroautophagy, microautophagy and chaperone-mediated autophagy), and the choice of which to use depends on how the proteins are identified and delivered to lysosomes12 (Fig. 1). The association of other chaperones and co-chaperones, such as CHIP or BAG3, with HSC70 determines HSC70-mediated targeting of proteins through degradation by the proteasome and macroautophagy, respectively. Properties in the cargo protein also contribute to the selection of the degradation pathway. For example, although single unfolded proteins can be eliminated through almost any degradation pathway, once multiple proteins organize into oligomeric complexes or aggregates, they can only be degraded by means of a selective form of macroautophagy known as aggrephagy13, or chaperone-assisted selective autophagy if mediated by HSC70 (ref. 14). The exposure of specific regions in the amino acid sequence of the protein or of degradation tags, acquired through posttranslational modification (i.e., ubiquitination, acetylation, etc.) also determine the degrada-tion pathway15.
Many age-related cellular changes can influence chaperoning activities. Thus, poor cellular energetics—a feature characteristic of old organisms owing to reduced mitochondrial function and dysregula-tion of lipid and glucose metabolism, etc.16,17—often limit the amount of available ATP. These differences in the bioavailability of ATP with age could be responsible for the repression of the ATP-dependent chaperones and the induction of ATP-independent chaperones that have been recently identified in the aging brain18. Similarly, the frac-tion of chaperones that is available for cargo recognition can also become markedly reduced with age. For example, sustained chronic stressors, such as the continued presence of a metastable protein, have been shown to act as a ‘sink’ for chaperones19. The loss of chaperone function and a reduction in availability further aggravate the problems with protein quality control. Undesired age-related modifications in
the substrate protein can also interfere with the chaperone’s ability to recognize its target. For example, the accumulation with aging of advanced glycation end-products through non-enzymatic modi-fications on long-lived proteins interferes with normal chaperone function. This type of modification is amenable to repair by enzymes such as methionine sulfoxide reductase, but the abundance of these enzymes decreases with age, further contributing to an accumulation of altered proteins that are unrecognizable to chaperones20. Cells may accommodate some of these age-related changes by modifying the pool of chaperones involved in proteostasis. Thus, although the HSP70 and HSP90 heat-shock protein families of chaperones have well-recognized roles in proteome balance under normal conditions, recent studies in nematodes highlight a prominent function of small heat-shock proteins in the preservation of proteostasis in aging; such small heat-shock proteins trap excess cytosolic proteins into protective aggregates21,22.
Autophagy and proteasome activity. Age-related changes in prote-ostasis are not restricted to chaperones. Autophagy and proteasome activity both decrease with aging3,23, but they do so to a lesser degree in association with healthy aging and longevity, such as in centenar-ians and long-lived animal species (i.e., the naked mole rat)7. Multiple types of interventions support the idea that diminishing the proteo-toxic load during aging can improve lifespan or healthspan (Table 1). Promoting proteasome or autophagy activity via the overexpres-sion of proteasome subunits or essential autophagy genes increases lifespan and confers resistance to stress in Saccharomyces cerevisiae, Caenorhabditis elegans and Drosophila melanogaster24,25. Information on such interventions in mammals is just beginning to emerge. For example, whole-body overexpression of the essential autophagy gene Atg5 in mice revealed anti-aging phenotypes and a lifespan exten-sion of about 20% (ref. 26). Interestingly, most of the interventions that slow down aging in experimental models are associated with improved proteostasis, and in many instances, these interventions
Exosome
Multivesicular body orlate endosome
Ribosome
Folded protein Misfolded protein
AggregateAutophagosome
Aggregates
Macroautophagy
HSC70
L2A
Proteasome
Chaperones
Lysosome
Chaperonedepletion
MA activityCargo recognitionPhagophore inductionAPG-LYS fusion
CMA activityL2A levels
UPS activityUbiquitinationSubunit levelsAssembly
Proteinsynthesis
UPS
CMA
Figure 1 Changes with age in intracellular proteostasis systems. Chaperones and two proteolytic systems, the ubiquitin proteasome system (UPS) and autophagy, take care of maintenance of intracellular proteostasis. Chaperones (blue, yellow and gray circles) assist de novo synthesized proteins and unfolded proteins to reach their folded stable status. If folding is not possible, chaperones target the unfolded protein for degradation by the proteasome (often after ubiquitination) or in lysosomes. Single soluble proteins can reach the lysosomal lumen through a membrane transporter in chaperone-mediated autophagy (CMA). Once misfolded proteins organize into oligomers or insoluble aggregates, the only options for their elimination from the cytosol are either by degradation in lysosomes through macroautophagy (MA) or expulsion outside the cell by means of small vesicles (exosomes). Red boxes indicate changes with age in different steps or components of the intracellular proteostasis networks. APG-LYS, autophagosome-lysosome; HSC70, heat-shock cognate protein of 70kDa; L2A, lysosome-associated membrane protein type 2A.
1408 VOLUME 21 | NUMBER 12 | DECEMBER 2015 nature medicine
demonstrated autophagy-activating pro-perties. For example, calorie restriction, rapamycin, metformin, resveratrol and spermidine, which are well known for their ability to extend lifespan and/or health-span, have all been proven to directly activate autophagy, although probably through different mechanisms25. Because at least some of these interventions could also affect protein homeostasis through their effects on chaperone levels and protein syn-thesis, the contribution of the autophagy- activating properties to their overall influ-ence on lifespan is currently under study. Furthermore, protein degradation—and proteostasis in general—can also be positively influenced by known non-pharmacological inducers. The activating effect of physical exercise on the activity of chaperones, on the proteasome and on some types of autophagy has been well documented27–30. Similarly, dietary interventions can also positively influ-ence proteostasis in most cases, through the activation of protein degradation or through changes in the balance between protein syn-thesis and degradation. Thus, in addition to starvation—one of the best-known inducers of autophagy—common dietary components such as olive oil, vitamins and even coffee have also been shown to have stimulatory properties with respect to different protein quality systems31,32.
Compared with the many studies highlighting the possible anti-aging value of improving protein degradation, the potential of improv-ing protein repair has been poorly explored. One limitation to the latter approach is limited knowledge about repair enzymes and how they change with age. A recent study observed lifespan extension in flies that overexpressed methionine sulfoxide reductase, the enzyme that reduces oxidized methionine20; this provides new support for fur-ther exploring the anti-aging value that manipulating levels of these repair enzymes could have.
Interestingly, recent studies suggest that the functional decline of the proteostasis networks may occur earlier than anticipated in the lifespan of an organism. In fact, studies in C. elegans have challenged the idea that proteostasis failure results from the gradual accumula-tion of cellular anomalies, instead pointing to programmed events at an early age as being responsible for the proteostasis collapse later in life33. At present, such observations are limited to invertebrates. If this finding were to hold true in mammals, however, it would become important to identify this early time frame and to determine whether it is common for the whole organism or whether there are organ- specific time differences; understanding these factors would help to correctly tailor interventions that could ameliorate the loss of proteostasis with age or in age-related diseases.
Intracellular proteostasis: cytosolic and organelle protein quality controlCytosolic proteostasis. Most advances in our understanding of cellular proteostasis and its changes with age arose from the study of the cytosolic proteostasis system34,35. In these studies, the cytosolic proteome has
often been considered to be a uniform entity in which most proteins undergo similar changes. But is that true? Do all proteins have similar susceptibility to cellular aggressors? Are there subgroups of proteins whose members are more prone to losing their stable conformations than are others? Potentially, there are also differences in the time-course in which different proteins escape homeostasis. Recent studies in C. elegans have analyzed these and related questions by monitoring more than 5,000 proteins during the worm lifespan. Contrary to predictions, the proteome changes with age were far from ‘subtle’, although they were gradual. A pronounced increase in protein abundance seems to be the main reason behind proteome imbalance, loss of stoichiometry and protein aggregation22. Similar future studies on the aging mammalian proteome should help to clarify the universality of the central role of changes in protein abundance in the loss of proteostasis with age.
One of the recent advances in our understanding of cytosolic prote-ostasis is the realization that proteolytic systems function in a coordi-nated manner. Proteasome and autophagy share substrates, effectors and regulators, which allows for continuous cross talk between these pathways and compensation for one another36. This compensation is advantageous for cells in disease conditions or in aging when one of the proteolytic systems ceases to work properly. For example, a grow-ing number of reports illustrates how a loss of proteostasis resulting from blockage of the proteasome is prevented by an upregulation of macroautophagy37. Similarly, cells react to the inhibition of one form of autophagy by activating a different one38,39. These pathways are never redundant, and the loss of one of them becomes evident once cells are exposed to stress; however, at least under basal conditions, these compensatory activities are able to preserve homeostasis36.
Table 1 Examples of interventions that increase lifespan by reducing proteotoxicityProteostasis system Organism Genetic or pharmacologic intervention Reference
Chaperones C. elegans HSP16 overexpression 91
C. elegans CR 92
Protein repair enzymes D. melanogaster Protein repair methyltransferase overexpression 93
D. melanogaster Methionine sulfoxide reductase overexpression 94
Ubiquitin-proteasome system
S. cerevisiae Loss of Ubr2 (negative regulator of UPS transcription) 95
C. elegans Rpn6 overexpression 96
C. elegans CR 97
D. melanogaster FOXO overexpression 98
D. melanogaster Rpn11 overexpression 99
M. musculus IGF1 overexpression 100
M. musculus CR 24
Autophagy C. elegans IGF signaling loss 101
C. elegans CR 102
C. elegans Resveratrol 103
C. elegans Spermidine 104
D. melanogaster ATG8a overexpression 105
D. melanogaster Spermidine 104
D. melanogaster Rapamycin 106
M. musculus ATG5 overexpression 26
Organelle S. cerevisiae Deletion of UPR genes 107
S. cerevisiae CR 108
S. cerevisiae CR 109
C. elegans Increased XBP-1 and IRE-1 in daf-2 mutants 110
C. elegans Activated ER UPR and mtUPR 111
C. elegans CR 112
D. melanogaster CR 113
D. melanogaster Hsp22 overexpression 114
D. melanogaster Overexpression of mtUPR genes 115
Abbreviations: CR, caloric restriction; ER, endoplasmic reticulum; UPR, unfolded protein response; mtUPR, mitochondrial UPR
nature medicine VOLUME 21 | NUMBER 12 | DECEMBER 2015 1409
Such coordinated functioning of the protein-degradation pathways should be kept in mind when interpreting results from the experimen-tal inhibition of one of these pathways. Most of this compensation has been documented in cultured cells, but recent studies support its occurrence in whole organisms as well. Autophagy is compensatorily activated in the rat hippocampus in response to proteasomal inhibi-tion40; similarly, mice in which chaperone-mediated autophagy in the liver has been blocked display a robust activation of macroautophagy and the proteasome system, both of which contribute to proteostasis in this organ41. However, compensation among proteolytic systems seems to be organ-dependent. For example, this compensation of chaperone-mediated autophagy failure by macroautophagy described in the liver is not observed in the retina42. In addition, for reasons still unknown, aging has a negative effect on the cross talk between proteolytic pathways. For example, in aged mice, the compensatory upregulation of some of these proteolytic pathways is lost40,41. The failure with age of signaling pathways such as the insulin-like growth factor 1 (IGF1) pathway (in the case of proteasome-autophagy cross talk)40, or of the transcription factor EB (TFEB) pathway (in cross talk between macroautophagy and chaperone-mediated autophagy)41, have been implicated in the loss of these compensatory abilities in aged organisms. Interestingly, chronic, moderate blockage of some subunits of the proteasome improves the ability of this system to respond to an acute proteotoxic insult43. This rebalancing of prote-ostasis in response to moderate blockage is conserved from yeast to humans43, but the underlying mechanisms that enable the cross talk and the contribution of the cross talk to these changes need further investigation.
Organelle proteostasis. Despite the empha-sis of a vast part of the literature on cytosolic proteostasis, the importance of protein home-ostasis inside organelles and the existence of organelle-specific proteostasis mechanisms are now well accepted (Fig. 2). The high volume of proteins—mostly in their native
states—handled by the endoplasmic reticulum (ER) creates a need for robust systems of protein-quality surveillance in this organelle44. A three-arm transcriptional program known as the unfolding protein response (UPR)—activated in response to a loss of proteostasis in the ER—and a dedicated retrotranslocation system, which allows for the rerouting of unfolded luminal and membrane proteins toward the cytosol for degradation, together ensure ER homeostasis44. The UPR is initiated by three ER transmembrane proteins, IRE1, PERK and ATF6, which subsequently induce the expression of several genes involved in protein processing and maturation in order to counteract the ER stress and to restore the proteostatic balance (Fig. 2). If the amount of unfolded proteins in the ER exceeds the ability of the UPR to return the ER proteome to homeostasis, retrotranslocation is acti-vated and this results in the arrival of the unfolded protein pool into the cytosol45. The proteasome was thought to be the only degrada-tion pathway involved in this process, but recent studies support the importance of a selective form of autophagy, now termed ER-phagy, for the maintenance of ER homeostasis. In this case, degradation includes not only unfolded proteins but also whole regions of the ER-limiting membrane, where the ER-resident protein FAM134B local-izes and acts as a receptor to facilitate the degradation of the ER46.
The tight connections between ER homeostasis and these two cytosolic proteolytic systems make the ER vulnerable to age-related changes in the activity of these systems. In addition, sustained ER stress and/or inappropriate response to this stress are common characteristics of aging and of many chronic age-related disorders (Table 2). This justifies recent efforts to chemically attenuate such
Proteasome
Lysosome
Autophagosome
Autophagosome
Endoplasmic reticulum
Phagophore
Phagophore
Mitochondrion
ER-phagy
Mitophagy
mtUPR
UPR
ERAD
Nucleus
FAM134B
PERK
IRE1
ATF6 XBP1
ATF4
ClpXPprotease
Mitochondrialchaperones
ERchaperones
Ubiquitin
Figure 2 Organelle proteostasis networks. Schematic of the mechanisms that preserve proteostasis in the endoplasmic reticulum (ER) and in mitochondria. The ER and mitochondria can undergo degradation as a whole organelle through specialized forms of autophagy, ER-phagy (assisted by the recently identified FAM134B protein) and mitophagy, respectively. In addition, these organelles have their own proteostasis systems that are activated by the presence of unfolded proteins. In the ER, these unfolded proteins activate the unfolded protein response (UPR) that has three arms (mediated by PERK and ATF4, IRE1 and XBP1, and by ATF6). Activation of the UPR attenuates protein translation and enhances the expression of ER chaperones. If this response is not sufficient, unfolded proteins are retrotranslocated for degradation in the cytoplasm by the proteasome (ERAD). The mitochondrial UPR (mtUPR) is similarly activated in response to proteotoxic stress to enhance chaperone content in mitochondria and to attenuate translation. Unfolded mitochondrial proteins are cleaved by the ClpXP protease into small peptides that upon translocation into the cytosol activate the mtUPR. D
1410 VOLUME 21 | NUMBER 12 | DECEMBER 2015 nature medicine
sustained ER stress. Most of the small molecules that are currently under development or in testing for the treatment of age-related disorders such as neurodegeneration or metabolic disorders target components of the UPR47,48. However, recent findings in worms have revealed that the UPR can also be activated by lipid disturbances alone (without disturbed proteostasis)49, opening the tantalizing possibility of using dietary changes to modulate this response in situations of ER proteo-toxicity. Under these conditions, stimulation of autophagy could also be valuable for alleviating the overloaded stress. Until now, however, most compounds undergoing testing have resulted in global activation of the autophagy process rather than the more desirable selective
enhancement of ER-phagy. Given that overexpression of FAM134B causes ER fragmentation and lysosomal degradation46, FAM134B could become a suitable target of specific ER-phagy activators.
The impact of aging on recently identified ER proteostasis mecha-nisms remains largely unexplored. For example, ER proteostasis can be induced—even before the UPR is activated—via the delivery of misfolded proteins to the cell surface, after which they can be inter-nalized and degraded in lysosomes50. A better understanding of the molecular effectors of this rapid ER stress–induced export (RESET) could provide additional targets to alleviate the loss of ER proteostasis in aging and some of the chronic age-related diseases.
Table 2 Changes in components of the proteostasis networks with age and in some age-related diseasesProteostasis system Change Aging condition or age-related disease
nature medicine VOLUME 21 | NUMBER 12 | DECEMBER 2015 1411
Remarkable progress has also been made in understanding the mechanisms behind mitochondrial proteostasis (Fig. 2). Maintenance of mitochondrial proteostasis is particularly challenging because of the abundance of reactive oxygen species (ROS) in this compartment. This probably explains why damaged portions of the mitochondrial network that succumb to severe proteotoxicity are often segregated from the network and eliminated as a whole organelle. The most com-mon way of getting rid of malfunctioning mitochondria is through autophagic degradation (mitophagy)51. The molecular complexity of this process, the variety of mitophagy types that coexist in the cell and the recently identified alternative ways by which the lysosomal system can contribute to mitochondria degradation independent of the autophagy machinery51 all highlight our relatively limited knowledge about mitochondrial clearance from the cell. However, numerous studies have reported the malfunctioning of mitophagy and thus mitochondrial clearance in aging and in age-related disorders (Table 2), in particular those involving neurodegeneration, making the study of these processes a high priority in these fields.
Although mitochondrial quality control is often tightly linked to the proteolytic cytosolic systems, this organelle, similarly to the ER, con-tains its own set of chaperones and proteases. The mitochondrial UPR (mtUPR) is activated when unfolded or misfolded proteins accumulate in the mitochondria and aggregate52,53. Recent studies have shown that communication of the mtUPR with the nucleus can induce the expres-sion of nuclear genes encoding for mitochondrial proteins in response to impaired proteostasis54,55. The molecular players involved in mtUPR are the subjects of intensive investigation in aging research because the modulation of the mtUPR has been shown to have lifespan-extension properties in multiple organisms56. Some of the proteins connected with the mtUPR, such as the mitochondrial deacetylase SIRT3, are also well-known lifespan modulators whose deficiencies have been linked to a higher incidence of age-related diseases (neurodegen-eration, metabolic syndrome and cancer) (Table 2)57. Furthermore, age-related changes in some of the recently identified components of the mtUPR have also been described. For example, the expression of SIRT7, a component of the regulatory branch of the mtUPR, is reduced in aged hematopoietic stem cells, and its overexpression in this setting improved the regenerative capability of these cells58.
Quality control in the nuclear compartment is not as well understood. Historically, the nucleus has been viewed as less vul-nerable to proteotoxicity than other organelles because the complex nuclear envelope and tightly regulated nuclear transport systems limit contact between nuclear material and the crowded pro-aggregating cytoplasm milieu. However, it is now well accepted that chaperones, ubiquitin ligases and the proteasome present in the nucleoplasm all contribute to nuclear proteostasis59.
Interestingly, some of the proteins that contribute to nuclear pro-tein quality reside in the nucleus whereas others are shuttled from the cytosol or ER into the nucleus in times of proteotoxic stress60. This last group transiently co-localizes to nuclear aggregates of mutant huntingtin, ataxin-1 or TAR DNA-binding protein 43 (TDP43)— proteins associated, respectively, with Huntington’s disease, spinoc-erebellar ataxia 1 and amyotrophic lateral sclerosis61. This transport of chaperones to the nucleus in times of stress points to a sensing mechanism that generates a signal to induce entry into the nucleus of protein quality components to assist in restoring proteostasis. It is possible that this signal also attracts modifiers that tag proteins for degradation because certain ubiquitin ligases, such as UHRF2 and PML4, are known to recognize mutant polyglutamine proteins localized to the nucleus. What the nuclear sensors of proteotoxicity
are and how these pathways work remain to be elucidated. Also poorly understood is the occurrence of protein breakdown in the nucleo-plasm. Even though the proteasome has been shown to be responsible for the degradation of oxidatively damaged histones and of surplus ribosomal proteins62,63, it is not clear whether that degradation occurs in the nucleus or only once these proteins have been transported to the cytosol. In the case of pathogenic proteins that aggregate in the nucleoplasm, there is strong evidence supporting proteasome degradation in situ64. Currently, little is known about the possible changes in nuclear proteostasis during aging. Are there subsets of nuclear proteins—as is the case in the cytosol—that are more vulnerable to a loss of proteostasis in aging? Do the sensing mecha-nisms of nuclear proteotoxicity fail with age? Could part of the loss of proteostasis with age in the cytoplasm originate from an increase in the spillover from the nucleus of altered proteins?
In this respect, little is known about the rules that govern communi-cation between the cytoplasm and organelle proteostasis networks or whether they change in old organisms. Recently, the single-stranded DNA-binding mitochondrial protein SSBP1 was shown to translocate to the nucleus in response to heat shock to modulate the heat shock factor protein 1 (HSF1)-dependent expression of cytosolic, nuclear and mitochondrial chaperones—the first known example of intercompart-mental regulation of proteostasis65. We still do not know whether there is a hierarchical order whereby preservation of proteostasis in some compartments is prioritized over others, or whether poorly handled proteotoxicity in one cellular compartment is always systematically fed into another in a joint attempt to restore overall cellular prote-ostasis. Additionally, mechanisms that preserve quality control and proteostasis in other cellular compartments have been poorly explored. Turnover of whole organelles, rather than the selective degradation of only specific organelle–resident proteins, may be favored for cellular compartments such as the Golgi, endosomes or even lysosomes, for which full sequestration and degradation through a form of selective autophagy (lysophagy) has been described66. The dependence of these processes on autophagic effectors may make them subject to the same age-dependent malfunctioning described for autophagy23.
Intercellular proteostasis: integrating organ and tissue responsesMost studies to date have approached the analysis of age-related changes in proteostasis as a cell-autonomous problem. However, growing evidence supports the existence of regulatory, intercellular proteostasis networks that help in the coordinated response of tissues and organs to proteotoxic insults.
Whereas the effectors of intercellular proteostasis, chaperones and proteolytic systems are, for the most part, the same ones described for intracellular proteostasis, the key question for coordinated proteosta-sis between cells is how signals of proteostasis stress are transmitted from one cell to another. Some of the communication channels may be the same ones normally used by cells for intercellular communi-cation. For example, the coupling of intercellular gap junctions to the activation of autophagy has been demonstrated by showing that connexins—the main components of gap junctions—also function as endogenous inhibitors of autophagy67.
However, intercellular proteostasis is not limited to the transmission of stress signals across cells to boost activation of their intrinsic proteostasis machinery. Components of the proteostasis network, and even toxic proteins, can be transferred from one cell to another with the ultimate goal of preserving whole-organ proteostasis (Fig. 3). For example, studies in D. melanogaster expressing aggregation- prone proteins found that experimentally increasing chaperones
1412 VOLUME 21 | NUMBER 12 | DECEMBER 2015 nature medicine
(Hsp70 and Hsp40) in a group of cells improved proteostasis in other cells in the same tissue, thereby demonstrating non-cell-autonomous maintenance of proteostasis68.
Exosomes have emerged as important organelles in cellular intercommunication69. Originating from the invagination of the endosomal membrane, these small vesicles trap samples of the cytosol that are then released to the extracellular environment upon the fusion of multivesicular endosomes with the plasma membrane. Exosomes can act as vehicles for the exchange of chaperones and maybe for other proteostasis effectors (Fig. 3). For example, chaperones can be detected in exosomes; their abundance increases under conditions of proteotoxicity; and adding chaperone-containing exosomes to cul-tured cells expressing aggregation-prone proteins decreases inclusion body formation, demonstrating that exosomes can be an efficient mechanism for the transfer of chaperones between cells68.
It would be interesting to evaluate how the recently described decline of multivesicular endosomal proteostasis with age70 could affect the transferring abilities of exosomes in old organisms. In this respect, recent studies have compared neuronally derived exosomes from individuals diagnosed with Alzheimer’s disease before the onset of symptoms and up to 10 years after diagnosis with those from non-affected individuals. Results showed consistently higher exosomal levels of ubiquitinated proteins and lysosomal proteins, but lower abundance of HSC70 in the exosomes of those with Alzheimer’s disease71. Studies such as this one highlight the possible value of exosomal proteins as biomarkers of disease spreading, but they also reinforce the need for a better understanding of exosome biology as a way to elucidate changes in proteostasis across cells with age and in age-related diseases.
Another mechanism of the intercellular transfer of materials that is gaining considerable attention is the use of tunneling nanotubes that allow for the direct exchange of cellular components between non-adjacent cells that are separated by distances greater than 100 µm. The material exchanged through nanotubes ranges from regulatory miRNAs to whole organelles, such as lysosomes, mitochondria, endo-somes and lipid droplets (Fig. 3)72–74. Because tunneling nanotubes between neural stem cells and brain microvascular endothelial cells have been shown to provide neuroprotection75, it is appealing to think that the transfer of, for example, lysosomes from one cell to another could help to sustain high levels of autophagy without the need for lysosomal biogenesis in the stressed cell.
Besides the transfer of components of the proteostasis networks across cells, the idea that pathogenic proteins could transfer across cells in a prion-like manner has gained momentum as a possible model for the propagation of neurodegenerative diseases. In this model, aggregated proteins are released from the affected cells and travel to nearby healthy cells where they act as seeds for aggregate formation (Fig. 3). Several aggregation-prone proteins involved in neurodegenerative diseases (α-synuclein, tau, β-amyloid and super-oxide dismutase 1) have been detected in exosomes76,77. Consequently, exosomes could have tremendous potential as biomarkers for the diagnosis and prognosis of these diseases. Whether the spreading of protein aggregates via exosomes is a primary feature in these diseases or a ‘way out’ for aggregated proteins that are secondary to blockage
of intracellular proteostasis mechanisms—a common development in these diseases—requires future investigation. It is still not known whether this mechanism of propagation increases in physiological aging and contributes to the whole organ’s disturbance of proteostasis, but the role of prion-like propagation in neurodegenerative diseases of aging, including in Alzheimer’s disease and other tauopathies, as well as in Parkinson’s, Creutzfeldt-Jakob and Lou Gehrig’s diseases, as well as in tauopathies, is gaining acceptance. Because the majority of these neurodegenerative diseases are sporadic and late in onset, it has been proposed that prion accumulation occurs throughout life, and once it exceeds some critical threshold, prion spreading triggers global neurological dysfunction78. Although still not explored in detail, it is also possible that nanotubes participate in the spreading of protein aggregates; they have been implicated in transferring exogenous and endogenous prions between infected and naïve neuronal cells, and, in Creutzfeldt-Jakob disease79, from bone marrow–derived dendritic cells in the periphery to primary neurons.
Overall, this transfer of chaperones, proteases and protein aggregates across cells may result from the heterogeneity of the robustness of the proteostasis systems inside organs. Cell-type and regional differences in the abundance and activity of the proteasome or the autophagy-lysosomal system have been documented in different organs and have been correlated with variances in their proteostasis capabilities. These regional differences in proteostasis could determine regional susceptibility to disease. For example, the activity of chaperone-mediated autophagy, a type of autophagy shown to contribute to the degradation of α-synuclein, has been found to be lower in aggregation- prone regions of the brain in both control animals and animals expressing the pathogenic form of α-synuclein80. In this context, intercellular proteostasis could initially serve to sustain proteosta-sis in the weakest regions by transferring aggregates out of the brain region toward regions with a more robust proteostasis and trafficking the components of the proteostasis networks in the reverse direction (Fig. 3). The growing repertoire of fluorescent and radiometric probes available for the study of different proteostasis events (protein folding, proteasome and autophagy activities, the UPR response, etc.) in real time, and the newly developed single-cell image tracking systems81,82 should allow researchers to perform detailed time-course analysis of
ChaperonesProteasome Lysosomes
Exosomes
Nanotubes
Weakproteostasis
networks
Strongproteostasis
networks
Proteinaggregates Amyloid
Figure 3 Schematic of possible mediators of intercellular proteostasis: exosomes and nanotubes. It is feasible that bidirectional transfer allows for the replenishment of proteostasis effectors from the cell with more robust proteostasis to one with weaker proteostasis, and transport of protein aggregates and damaged proteins in the reverse direction.
nature medicine VOLUME 21 | NUMBER 12 | DECEMBER 2015 1413
aggregopathies in the near future. This type of analysis could help to identify tissue areas of proteostasis weakness, possible changes with age in the proteostatic strength of these areas and the contribution of nearby organ regions with better proteostasis to their response to toxic insults. Also pending is a better understanding of the impact that aging could have on the mechanisms that mediate intercellular prote-ostasis. Do transferring mechanisms, kinetics of transference or the amount of chaperones and proteases transferred between cells change with age? Do aging cells frequently use outside ‘dumping’ of protein aggregates in response to overloading of their intrinsic proteostasis machinery? Could exosome-based transfer be effective in boosting proteostasis in old organs?
Tele-proteostasis: integrating distant proteostasis networksThe realization that proteostasis networks in different organs coordinate with each other and that changes in one of them affect the functioning of the others has been one of the most exciting developments in the field of proteostasis in recent years. This ‘tele-proteostasis’ (as we term proteostasis coordinated from a distance through a combination of cell autonomous and non-autonomous mechanisms) allows, for example, for the induction of a systemic heat-shock response by activating the heat-shock protein response in neuronal cells alone62.
One of the first observations of interorgan regulation of proteos-tasis came from studies in worms expressing, in an inducible man-ner, proteins prone to misfolding that trigger the expression of the HSP90 chaperone to assist with protein refolding. Interestingly, such HSP90 expression was observed not only in the cells expressing the misfolded protein—in that case, muscle cells—but also in cells that did not express the misfolded protein83. In support of the idea that this distal proteostasis response was of value for handling protein misfolding in the affected cells, expression of HSP90 when restricted to neuronal or intestinal cells was still effective in resolving the aggre-gation problems in muscle cells83. These studies provided experimen-tal support that modulating the proteostasis responses in one organ elicits a similar response in a distant tissue83.
Shortly afterwards, it was demonstrated that tele-proteostasis applies to both cytosolic and organelle proteostasis, and also, that the robustness of tele-proteostasis is an important determinant of longevity. In worms that have been genetically modified to have a diminished systemic UPR (proven to be short lived), restoration of the missing UPR component in neurons and intestinal cells only was sufficient to increase the worms’ longevity84. In fact, neuronal res-toration of the missing UPR component activated the UPR in distal intestinal cells and promoted ER stress resistance in aged worms.
Although most of the discoveries in non-cell-autonomous prote-ostasis have been made in invertebrates, evidence of a similar proc-ess in mammals is starting to emerge. Constitutive expression of a component of the UPR in the hypothalamic neurons in mice also activated expression of this component in the liver, supporting cou-pling between ER stress in neurons and hepatocytes85. In this case, the consequences of such tele-proteostasis go beyond the maintenance of proteome stability in the distant organ; they also include important changes in hepatic metabolism that resonate in organismal energet-ics. Thus, ER stress in hypothalamic neurons is coupled to increased hepatic insulin sensitivity and also to suppressed glucose production in this organ85.
Since the discovery of this cell non-autonomous activation of the chaperone response and of the UPR and their links to longevity, a race has begun to identify the systemic mediators involved. UNC-13,
a protein that mediates neurotransmitter release in neurons, was one of the first components of neuroendocrine signaling to be implicated in distant UPR activation in worms84. As expected given the role of such a mediator, UNC-13 mutant worms also displayed reduced longevity. Secretion of a UPR component, the ER chaperone ERDJ3, has also been shown to contribute to extracellular proteos-tasis86, although whether or not this chaperone could also influ-ence proteostasis in distant tissues requires further investigation. Interestingly, the search for hormone-like factors that mediate proteostasis at a distance comes at a time when the aging research community is fully invested in the search for serum circulating factors with anti-aging properties. The concept originated from early studies showing the rejuvenating effect—at the stem cell level, at least—of administering serum from young mice to aged mice87 and has been recently extended as a protective mechanism against age-related neu-rodegeneration88. Now that some of the circulating factors respon-sible for these beneficial properties have been identified, a burning question is whether part of their positive effect is exerted through the regulation of proteostasis and whether they could be some of the sought-after mediators of tele-proteostasis. Furthermore, caloric restriction, an effective anti-aging intervention, has been extensively reported to improve proteostasis, and serum from caloric-restricted animals is known to delay senescence and improve stress resistance of cultured cells89. Both of these facts support the idea that circulat-ing factors under these conditions contribute to the regulation of multiple-organ proteostasis. Whether the circulating mediators of tele-proteostasis are universal and independent of the cellular condi-tions, or whether specific subsets of factors are used in response to different insults, however, requires further investigation.
Given that this is the most newly understood aspect of proteosta-sis, many additional interesting questions remain under the general umbrella of factors in the circulation that influence aging. Is there a role in interorgan proteostasis for inflammatory molecules, which are produced in abundance in chronic inflammatory processes in aging? Old organisms also undergo major metabolic changes; do circulating nutrients influence the coordination of proteostatic networks in dif-ferent organs? And of course, the question that we all are considering: can manipulation of proteostasis in one organ be used for therapeutic purposes in a distant one? It would be remarkable if some of the severe neurodegenerative diseases that affect the elderly could be treated with agents that target proteostasis in peripheral organs.
ConclusionsOverwhelming evidence supports the maintenance of cellular pro-teostasis as one of the key processes in ensuring longevity. A better understanding in recent years of the intracellular systems that contrib-ute to protein quality control has formed the conceptual framework for several successful attempts to improve cellular proteostasis to delay signs of aging and the progression of age-related diseases in some experimental models. Although many proof-of-principle interventions have so far relied on genetic manipulations, which are of questionable applicability in elderly patients, there are ongoing investigations using chemical targeting of chaperones or the proteolytic systems in the con-text of adult-onset neurodegeneration90. However, the pharmacopeia available for proteostasis modulation is still limited. The identification of new types of proteostasis (intercellular and interorgan) should help to expand the number of possible targets with therapeutic potential. Furthermore, consideration should be given to the effects of non-pharmacological interventions that are known to affect aging, such as exercise, diet, social interactions, behavior modifiers, etc.
1414 VOLUME 21 | NUMBER 12 | DECEMBER 2015 nature medicine
As a rapidly evolving field, proteostasis is in need of technological advances that allow for better read-outs. Different probes are now available for studying separate proteostasis events, but to gain an integrated understanding of organism proteostasis and its changes with age, we need methods that enable real-time and organ-spanning monitoring of proteostasis in whole organisms. Beyond technological advances, the other important change—fortunately, already under way—is a conceptual adjustment to the idea that proteostasis is not a cellular problem and that ‘it takes an organism’ to maintain it.
COMPETING FINANCIAL INTERESTSThe authors declare no competing financial interests.
Reprints and permissions information is available online at http://www.nature.com/reprints/index.html.
1. Vilchez, D., Saez, I. & Dillin, A. The role of protein clearance mechanisms in organismal ageing and age-related diseases. Nat. Commun. 5, 5659 (2014).
2. Roth, D.M. & Balch, W.E. Modeling general proteostasis: proteome balance in health and disease. Curr. Opin. Cell Biol. 23, 126–134 (2011).
3. Morimoto, R.I. & Cuervo, A.M. Proteostasis and the aging proteome in health and disease. J. Gerontol. A Biol. Sci. Med. Sci. 69 (suppl. 1), S33–S38 (2014).
4. Labbadia, J. & Morimoto, R.I. The biology of proteostasis in aging and disease. Annu. Rev. Biochem. 84, 435–464 (2015).
5. Labbadia, J. & Morimoto, R.I. Proteostasis and longevity: when does aging really begin? F1000Prime Rep. 6, 7 (2014).
6. Treaster, S.B. et al. Superior proteome stability in the longest lived animal. Age (Dordr) 36, 9597 (2014).
7. Pérez, V.I. et al. Protein stability and resistance to oxidative stress are determinants of longevity in the longest-living rodent, the naked mole-rat. Proc. Natl. Acad. Sci. USA 106, 3059–3064 (2009).
8. Tyedmers, J., Mogk, A. & Bukau, B. Cellular strategies for controlling protein aggregation. Nat. Rev. Mol. Cell Biol. 11, 777–788 (2010).
9. Feldman, D.E. & Frydman, J. Protein folding in vivo: the importance of molecular chaperones. Curr. Opin. Struct. Biol. 10, 26–33 (2000).
10. Navon, A. & Ciechanover, A. The 26 S proteasome: from basic mechanisms to drug targeting. J. Biol. Chem. 284, 33713–33718 (2009).
11. Tanaka, K. The proteasome: overview of structure and functions. Proc. Jpn. Acad. Ser. B Phys. Biol. Sci. 85, 12–36 (2009).
12. Kaushik, S. & Cuervo, A.M. Chaperones in autophagy. Pharmacol. Res. 66, 484–493 (2012).
13. Lamark, T. & Johansen, T. Aggrephagy: selective disposal of protein aggregates by macroautophagy. Int. J. Cell Biol. 2012, 736905 (2012).
14. Arndt, V. et al. Chaperone-assisted selective autophagy is essential for muscle maintenance. Curr. Biol. 20, 143–148 (2010).
15. Stolz, A., Ernst, A. & Dikic, I. Cargo recognition and trafficking in selective autophagy. Nat. Cell Biol. 16, 495–501 (2014).
16. Ma, Y. & Li, J. Metabolic shifts during aging and pathology. Compr. Physiol. 5, 667–686 (2015).
17. Ritz, P. & Berrut, G. Mitochondrial function, energy expenditure, aging and insulin resistance. Diabetes Metab. 31 (spec. no. 2), 5S67–5S73 (2005).
18. Brehme, M. et al. A chaperome subnetwork safeguards proteostasis in aging and neurodegenerative disease. Cell Rep. 9, 1135–1150 (2014).
19. Yu, A. et al. Protein aggregation can inhibit clathrin-mediated endocytosis by chaperone competition. Proc. Natl. Acad. Sci. USA 111, E1481–E1490 (2014).
20. Vanhooren, V. et al. Protein modification and maintenance systems as biomarkers of ageing. Mech. Ageing Dev. doi:10.1016/j.mad.2015.03.009 (2015).
21. Morrow, G., Samson, M., Michaud, S. & Tanguay, R.M. Overexpression of the small mitochondrial Hsp22 extends Drosophila life span and increases resistance to oxidative stress. FASEB J. 18, 598–599 (2004).
22. Walther, D.M. et al. Widespread proteome remodeling and aggregation in aging C. elegans. Cell 161, 919–932 (2015).
23. Rubinsztein, D.C., Marino, G. & Kroemer, G. Autophagy and aging. Cell 146, 682–695 (2011).
24. Chondrogianni, N., Georgila, K., Kourtis, N., Tavernarakis, N. & Gonos, E.S. 20S proteasome activation promotes life span extension and resistance to proteotoxicity in Caenorhabditis elegans. FASEB J. 29, 611–622 (2015).
25. Madeo, F., Zimmermann, A., Maiuri, M.C. & Kroemer, G. Essential role for autophagy in life span extension. J. Clin. Invest. 125, 85–93 (2015).
26. Pyo, J.O. et al. Overexpression of Atg5 in mice activates autophagy and extends lifespan. Nat. Commun. 4, 2300 (2013).
27. Morton, J.P., Kayani, A.C., McArdle, A. & Drust, B. The exercise-induced stress response of skeletal muscle, with specific emphasis on humans. Sports Med. 39, 643–662 (2009).
28. Ulbricht, A. et al. Induction and adaptation of chaperone-assisted selective autophagy CASA in response to resistance exercise in human skeletal muscle. Autophagy 11, 538–546 (2015).
29. He, C. et al. Exercise-induced BCL2-regulated autophagy is required for muscle glucose homeostasis. Nature 481, 511–515 (2012).
30. Jamart, C. et al. Modulation of autophagy and ubiquitin-proteasome pathways during ultra-endurance running. J. Appl. Physiol. 112, 1529–1537 (2012).
31. Katsiki, M., Chondrogianni, N., Chinou, I., Rivett, A.J. & Gonos, E.S. The olive constituent oleuropein exhibits proteasome stimulatory properties in vitro and confers life span extension of human embryonic fibroblasts. Rejuvenation Res. 10, 157–172 (2007).
32. Salomone, F. et al. Coffee enhances the expression of chaperones and antioxidant proteins in rats with nonalcoholic fatty liver disease. Transl. Res. 163, 593–602 (2014).
33. Ben-Zvi, A., Miller, E.A. & Morimoto, R.I. Collapse of proteostasis represents an early molecular event in Caenorhabditis elegans aging. Proc. Natl. Acad. Sci. USA 106, 14914–14919 (2009).
34. Iram, A. & Naeem, A. Protein folding, misfolding, aggregation and their implications in human diseases: discovering therapeutic ways to amyloid-associated diseases. Cell Biochem. Biophys. 70, 51–61 (2014).
35. Kim, Y.E., Hipp, M.S., Bracher, A., Hayer-Hartl, M. & Hartl, F.U. Molecular chaperone functions in protein folding and proteostasis. Annu. Rev. Biochem. 82, 323–355 (2013).
36. Park, C. & Cuervo, A.M. Selective autophagy: talking with the UPS. Cell Biochem. Biophys. 67, 3–13 (2013).
37. Korolchuk, V.I., Menzies, F.M. & Rubinsztein, D.C. Mechanisms of cross-talk between the ubiquitin-proteasome and autophagy-lysosome systems. FEBS Lett. 584, 1393–1398 (2010).
38. Kaushik, S., Massey, A., Mizushima, N. & Cuervo, A.M. Constitutive activation of chaperone-mediated autophagy in cells with impaired macroautophagy. Mol. Biol. Cell 19, 2179–2192 (2008).
39. Massey, A.C., Kaushik, S., Sovak, G., Kiffin, R. & Cuervo, A.M. Consequences of the selective blockage of chaperone-mediated autophagy. Proc. Natl. Acad. Sci. USA 103, 5805–5810 (2006).
40. Gavilán, E. et al. Age-related dysfunctions of the autophagy lysosomal pathway in hippocampal pyramidal neurons under proteasome stress. Neurobiol. Aging 36, 1953–1963 (2015).
41. Schneider, J.L. et al. Loss of hepatic chaperone-mediated autophagy accelerates proteostasis failure in aging. Aging Cell 14, 249–264 (2015).
42. Rodríguez-Muela, N. et al. Balance between autophagic pathways preserves retinal homeostasis. Aging Cell 12, 478–488 (2013).
43. Tsvetkov, P. et al. Compromising the 19S proteasome complex protects cells from reduced flux through the proteasome. eLIFE 4 doi:10.7554/eLife.08467 (2015).
44. Walter, P. & Ron, D. The unfolded protein response: from stress pathway to homeostatic regulation. Science 334, 1081–1086 (2011).
45. Brodsky, J.L. Cleaning up: ER-associated degradation to the rescue. Cell 151, 1163–1167 (2012).
46. Khaminets, A. et al. Regulation of endoplasmic reticulum turnover by selective autophagy. Nature 522, 354–358 (2015).
47. Moreno, J.A. et al. Oral treatment targeting the unfolded protein response prevents neurodegeneration and clinical disease in prion-infected mice. Sci. Transl. Med. 5, 206ra138 (2013).
49. Hou, N.S. et al. Activation of the endoplasmic reticulum unfolded protein response by lipid disequilibrium without disturbed proteostasis in vivo. Proc. Natl. Acad. Sci. USA 111, E2271–E2280 (2014).
50. Satpute-Krishnan, P. et al. ER stress-induced clearance of misfolded GPI-anchored proteins via the secretory pathway. Cell 158, 522–533 (2014).
51. Lemasters, J.J. Variants of mitochondrial autophagy: Types 1 and 2 mitophagy and micromitophagy (Type 3). Redox Biol. 2, 749–754 (2014).
52. Heo, J.M. et al. A stress-responsive system for mitochondrial protein degradation. Mol. Cell 40, 465–480 (2010).
53. Braun, R.J. et al. Accumulation of basic amino acids at mitochondria dictates the cytotoxicity of aberrant ubiquitin. Cell Rep. 10, 1557–1571 (2015).
54. Jovaisaite, V. & Auwerx, J. The mitochondrial unfolded protein response-synchronizing genomes. Curr. Opin. Cell Biol. 33, 74–81 (2015).
55. Haynes, C.M. & Ron, D. The mitochondrial UPR - protecting organelle protein homeostasis. J. Cell Sci. 123, 3849–3855 (2010).
56. Jensen, M.B. & Jasper, H. Mitochondrial proteostasis in the control of aging and longevity. Cell Metab. 20, 214–225 (2014).
57. McDonnell, E., Peterson, B.S., Bomze, H.M. & Hirschey, M.D. SIRT3 regulates progression and development of diseases of aging. Trends Endocrinol. Metab. 26, 486–492 (2015).
58. Mohrin, M. et al. Stem cell aging. A mitochondrial UPR-mediated metabolic checkpoint regulates hematopoietic stem cell aging. Science 347, 1374–1377 (2015).
59. Shibata, Y. & Morimoto, R.I. How the nucleus copes with proteotoxic stress. Curr. Biol. 24, R463–R474 (2014).
60. Andersen, J.S. et al. Nucleolar proteome dynamics. Nature 433, 77–83 (2005).61. Janer, A. et al. PML clastosomes prevent nuclear accumulation of mutant ataxin-
7 and other polyglutamine proteins. J. Cell Biol. 174, 65–76 (2006).62. Ullrich, O. et al. Poly-ADP ribose polymerase activates nuclear proteasome to
degrade oxidatively damaged histones. Proc. Natl. Acad. Sci. USA 96, 6223–6228 (1999).
67. Bejarano, E. et al. Connexins modulate autophagosome biogenesis. Nat. Cell Biol. 16, 401–414 (2014).
68. Takeuchi, T. et al. Intercellular chaperone transmission via exosomes contributes to maintenance of protein homeostasis at the organismal level. Proc. Natl. Acad. Sci. USA 112, E2497–E2506 (2015).
69. Lo Cicero, A., Stahl, P.D. & Raposo, G. Extracellular vesicles shuffling intercellular messages: for good or for bad. Curr. Opin. Cell Biol. 35, 69–77 (2015).
71. Goetzl, E.J. et al. Altered lysosomal proteins in neural-derived plasma exosomes in preclinical Alzheimer disease. Neurology 85, 40–47 (2015).
72. Astanina, K., Koch, M., Jungst, C., Zumbusch, A. & Kiemer, A.K. Lipid droplets as a novel cargo of tunnelling nanotubes in endothelial cells. Sci. Rep. 5, 11453 (2015).
73. Burtey, A. et al. Intercellular transfer of transferrin receptor by a contact-, Rab8-dependent mechanism involving tunneling nanotubes. FASEB J. 29, 4695–4712 (2015).
74. Wang, X. & Gerdes, H.H. Transfer of mitochondria via tunneling nanotubes rescues apoptotic PC12 cells. Cell Death Differ. 22, 1181–1191 (2015).
75. Wang, X. et al. Rescue of brain function using tunneling nanotubes between neural stem cells and brain microvascular endothelial cells. Mol. Neurobiol. doi:10.1007/s12035-015-9225-z (2015).
76. Agosta, F., Weiler, M. & Filippi, M. Propagation of pathology through brain networks in neurodegenerative diseases: from molecules to clinical phenotypes. CNS Neurosci. Ther. 21, 754–767 (2015).
77. Russo, I., Bubacco, L. & Greggio, E. Exosomes-associated neurodegeneration and progression of Parkinson′s disease. Am. J. Neurodegener. Dis. 1, 217–225 (2012).
78. Prusiner, S.B. Biology and genetics of prions causing neurodegeneration. Annu. Rev. Genet. 47, 601–623 (2013).
79. Gousset, K. et al. Prions hijack tunnelling nanotubes for intercellular spread. Nat. Cell Biol. 11, 328–336 (2009).
80. Malkus, K.A. & Ischiropoulos, H. Regional deficiencies in chaperone-mediated autophagy underlie alpha-synuclein aggregation and neurodegeneration. Neurobiol. Dis. 46, 732–744 (2012).
81. Liu, Y., Zhang, X., Chen, W., Tan, Y.L. & Kelly, J.W. Fluorescence turn-on folding sensor to monitor proteome stress in live cells. J. Am. Chem. Soc. 137, 11303–11311 (2015).
82. Arrasate, M. & Finkbeiner, S. Automated microscope system for determining factors that predict neuronal fate. Proc. Natl. Acad. Sci. USA 102, 3840–3845 (2005).
83. van Oosten-Hawle, P., Porter, R.S. & Morimoto, R.I. Regulation of organismal proteostasis by transcellular chaperone signaling. Cell 153, 1366–1378 (2013).
84. Taylor, R.C. & Dillin, A. XBP-1 is a cell-nonautonomous regulator of stress resistance and longevity. Cell 153, 1435–1447 (2013).
85. Williams, K.W. et al. Xbp1s in Pomc neurons connects ER stress with energy balance and glucose homeostasis. Cell Metab. 20, 471–482 (2014).
86. Genereux, J.C. et al. Unfolded protein response-induced ERdj3 secretion links ER stress to extracellular proteostasis. EMBO J. 34, 4–19 (2015).
87. Conboy, M.J., Conboy, I.M. & Rando, T.A. Heterochronic parabiosis: historical perspective and methodological considerations for studies of aging and longevity. Aging Cell 12, 525–530 (2013).
88. Katsimpardi, L. et al. Vascular and neurogenic rejuvenation of the aging mouse brain by young systemic factors. Science 344, 630–634 (2014).
89. de Cabo, R. et al. Serum from calorie-restricted animals delays senescence and extends the lifespan of normal human fibroblasts in vitro. Aging (Albany, NY) 7, 152–166 (2015).
90. Pratt, W.B., Gestwicki, J.E., Osawa, Y. & Lieberman, A.P. Targeting Hsp90/Hsp70-based protein quality control for treatment of adult onset neurodegenerative diseases. Annu. Rev. Pharmacol. Toxicol. 55, 353–371 (2015).
91. Walker, G.A. & Lithgow, G.J. Lifespan extension in C. elegans by a molecular chaperone dependent upon insulin-like signals. Aging Cell 2, 131–139 (2003).
92. Steinkraus, K.A. et al. Dietary restriction suppresses proteotoxicity and enhances longevity by an hsf-1-dependent mechanism in Caenorhabditis elegans. Aging Cell 7, 394–404 (2008).
93. Chavous, D.A., Jackson, F.R. & O’Connor, C.M. Extension of the Drosophila lifespan by overexpression of a protein repair methyltransferase. Proc. Natl. Acad. Sci. USA 98, 14814–14818 (2001).
94. Ruan, H. et al. High-quality life extension by the enzyme peptide methionine sulfoxide reductase. Proc. Natl. Acad. Sci. USA 99, 2748–2753 (2002).
95. Kruegel, U. et al. Elevated proteasome capacity extends replicative lifespan in Saccharomyces cerevisiae. PLoS Genet. 7, e1002253 (2011).
96. Vilchez, D. et al. RPN-6 determines C. elegans longevity under proteotoxic stress conditions. Nature 489, 263–268 (2012).
97. Depuydt, G. et al. Reduced insulin/insulin-like growth factor-1 signaling and dietary restriction inhibit translation but preserve muscle mass in Caenorhabditis elegans. Mol. Cell. Proteomics 12, 3624–3639 (2013).
98. Demontis, F. & Perrimon, N. FOXO/4E-BP signaling in Drosophila muscles regulates organism-wide proteostasis during aging. Cell 143, 813–825 (2010).
99. Tonoki, A. et al. Genetic evidence linking age-dependent attenuation of the 26S proteasome with the aging process. Mol. Cell. Biol. 29, 1095–1106 (2009).
100. Crowe, E., Sell, C., Thomas, J.D., Johannes, G.J. & Torres, C. Activation of proteasome by insulin-like growth factor-I may enhance clearance of oxidized proteins in the brain. Mech. Ageing Dev. 130, 793–800 (2009).
101. Meléndez, A. et al. Autophagy genes are essential for dauer development and life-span extension in C. elegans. Science 301, 1387–1391 (2003).
102. Jia, K. & Levine, B. Autophagy is required for dietary restriction-mediated life span extension in C. elegans. Autophagy 3, 597–599 (2007).
103. Morselli, E. et al. Caloric restriction and resveratrol promote longevity through the Sirtuin-1-dependent induction of autophagy. Cell Death Dis. 1, e10 (2010).
104. Eisenberg, T. et al. Induction of autophagy by spermidine promotes longevity. Nat. Cell Biol. 11, 1305–1314 (2009).
105. Simonsen, A. et al. Promoting basal levels of autophagy in the nervous system enhances longevity and oxidant resistance in adult Drosophila. Autophagy 4, 176–184 (2008).
106. Bjedov, I. et al. Mechanisms of life span extension by rapamycin in the fruit fly Drosophila melanogaster. Cell Metab. 11, 35–46 (2010).
107. Labunskyy, V.M. et al. Lifespan extension conferred by endoplasmic reticulum secretory pathway deficiency requires induction of the unfolded protein response. PLoS Genet. 10, e1004019 (2014).
108. Barros, M.H., Bandy, B., Tahara, E.B. & Kowaltowski, A.J. Higher respiratory activity decreases mitochondrial reactive oxygen release and increases life span in Saccharomyces cerevisiae. J. Biol. Chem. 279, 49883–49888 (2004).
109. McCormick, M.A. et al. A comprehensive analysis of replicative lifespan in 4,698 single-gene deletion strains uncovers conserved mechanisms of aging. Cell Metab. 22, 895–906 (2015).
110. Henis-Korenblit, S. et al. Insulin/IGF-1 signaling mutants reprogram ER stress response regulators to promote longevity. Proc. Natl. Acad. Sci. USA 107, 9730–9735 (2010).
111. Shore, D.E., Carr, C.E. & Ruvkun, G. Induction of cytoprotective pathways is central to the extension of lifespan conferred by multiple longevity pathways. PLoS Genet. 8, e1002792 (2012).
112. Chen, D., Thomas, E.L. & Kapahi, P. HIF-1 modulates dietary restriction-mediated lifespan extension via IRE-1 in Caenorhabditis elegans. PLoS Genet. 5, e1000486 (2009).
113. Zid, B.M. et al. 4E-BP extends lifespan upon dietary restriction by enhancing mitochondrial activity in Drosophila. Cell 139, 149–160 (2009).
114. Kim, H.J., Morrow, G., Westwood, J.T., Michaud, S. & Tanguay, R.M. Gene expression profiling implicates OXPHOS complexes in lifespan extension of flies over-expressing a small mitochondrial chaperone, Hsp22. Exp. Gerontol. 45, 611–620 (2010).
115. Owusu-Ansah, E., Song, W. & Perrimon, N. Muscle mitohormesis promotes longevity via systemic repression of insulin signaling. Cell 155, 699–712 (2013).
