MICRO. 555 (555 Microbial Molecular Genetics)
Dr.Afaf Ibrahim ShehataBotany and Microbiology Department
King Saud University
MOLECULAR GENETICS
molecular basis of inheritance Genes ---> Enzymes ---> Metabolism (phenotype)
Central Dogma of Molecular Biology* DNA -transcription--> RNA -translation--> Protein
Student CD Activity - 13.2 Events Protein Synthesis: INFORMATION FLOW
What is a GENE = ? DNA is the genetic material... [ but what about, retroviruses, as
HIV & TMV, contain RNA ] - a discrete piece of deoxyribonucleic acid
- linear polymer of repeating nucleotide monomers nucleotides* --> A adenine, C cytosine
T thymidine, G guanine --> polynucleotide*
Understanding Genetics
INFORMATION PROCESSING & the CENTRAL DOGMA
- the letters of the genetic alphabet... are the nucleotides A, T, G, & C of DNA
- the unit of information is CODON = genetic 'word'
a triplet sequence of nucleotides 'CAT' in a polynucleotide
3 nucleotides = 1 codon (word) = 1 amino acid
- the definition of (codon) word = amino acid
- Size of Human Genome: ≈ 3,000,000,000 base pairs or 1.5b in single strand of DNA genes
≈ 500,000,000 possible codons (words or amino acids)
- average page your textbook = approx 850 words thus, human genome is equal to 590,000 pages
or 470 copies of bio text book reading at 3 bases/sec it would take you about
47.6 years @ 8h/d - 7d/w WOW... extreme nanotechnology
µ Mice & humans (indeed, most or all mammals including dogs, cats, rabbits, monkeys, & apes)
have roughly the same number of nucleotides in their genomes -- about 3 billion
bp.
Experimental Proof of DNA as Genetic Material... 1. Transformation Experiments of Fred Griffith... (1920's)
Streptococcus pneumoniae - pathogenic S strain & benign R transforming 'principle'* (converting R to S cells) is the genetic
element
2. Oswald Avery, Colin MacLeod, & Maclyn McCarty... (1940's) suggest the transforming substance* is DNA molecules, but...
3. Alfred Hershey & Martha Chase's* 1952 bacteriophage experiments
*... VIRAL REPLICATION* [ pic 1 phage infection & pic-2* & lytic/lysogenic ]
a genetically controlled biological activity (viral reproduction) they did novel experiment... 1st real use radioisotopes in biology*
CONCLUSION - DNA is genetic material because (32P) nucleic acid not (35S) protein guides* viral replication
Sumanas, Inc. animation - Life cycle of HIV virus
Replication of DNA... (Arthur Kornberg - 1959 Nobel - died 10/26/07)
copying of DNA into DNA is structurally obvious??? [figure*]
Patterns of Replication* = conservative, semi-conservative, & dispersive
Matt Meselson & Frank Stahl 1958 - experimental design* can we separate 15N-DNA from 14N-DNA - (OLD DNA from
NEW DNA)? sedimentation of DNA's (sucrose gradients -->
CsCl gradients* & picture*) we can predict results... figure* & overview & all possible
results Sumanas, Inc. animation - Meselson-Stahl
DNA Replications
DNA polymerase: enzyme that copies DNA... prokaryotic Pol I-IV eukaryotic a & d
Pol III (pic) req: 4-deoxy-NTP's & ssDNA template piece reads template and adds a complimentary nucleotide*
reads 3' to 5' and synthesizes in 5' to 3' direction...
[quicktime movie] proofreads* & bidirectional synthesis*... & EM pic*
Replication forks - leading & lagging strands - Campbell figure*
Arthur Kornberg - 1st to synthesize DNA in test tube, died 26 Oct 2007
Model of Replication is bacterial with DNA polymerase III...
several enzymes* form a Replication Complex (Replisome) & include:
helicase - untwists DNA topoisomerase [DNA gyrase] - removes supercoils,
single strand binding proteins - stabilize replication fork, Primase - makes RNA primer
POL III - synthesizes new DNA strands DNA polymerase I - removes RNA primer 1 base at a time,
adds DNA bases DNA ligase repairs Okazaki fragments (seals lagging strand 3'
open holes)
Structure of DNA polymerase III* copies both strands simultaneously, as DNA is
Threaded Through a Replisome* a "replication machine", which may be
stationary by anchoring in nuclear matrix Continuous & Discontinuous replication
occur simultaneously in both strands
EVENTS: 1. DNA pol III binds at the origin of replication site in the template strand 2. DNA is unwound by replisome complex using helicase & topoisomerase
3. all polymerases require a preexisting DNA strand (PRIMER) to start replication,
thus Primase adds a single short primer to the LEADING strand and adds many primers to the LAGGING strand
4. DNA pol III is a dimer adding new nucleotides to both strands primers direction of reading is 3' ---> 5' on template
direction of synthesis of new strand is 5" ---> 3' rate of synthesis is substantial 400 nucleotide/sec
5. DNA pol I removes primer at 5' end replacing with DNA bases, leaves 3' hole
6. DNA ligase seals 3' holes of Okazaki fragments on lagging strand the sequence and DNA Repair*
Rates of DNA synthesis: myDNAi movie of replication* native polymerase: 400 bases/sec with 1
error per 109 bases artificial: phophoramidite method
(Marvin Caruthers, U.Colorado); ssDNA synthesis on polystyrene bead @ 1 base/300 sec with
error rate of 1/100b
GENE Expression the Central Dogma of Molecular Biology depicts flow of genetic
information Transcription - copying of DNA sequence into RNA
Translation - copying of RNA sequence into protein
DNA sequence -------> RNA sequence -----> amino acid sequence
TAC AUG MET triplet sequence in DNA --> codon in mRNA ----> amino acid
in protein
Information : triplet sequence in DNA is the genetic word [codon]
Compare Events: Procaryotes* vs. Eucaryotes* = Separation of labor
Differences DNA vs. RNA (bases & sugars) and its single stranded
Flow of Gene Information (FIG*) - One Gene - One enzyme (Beadle & Tatum
Transcription - RNA polymerase RNA*polymerase - in bacteria Sigma factor* binds promoter
& initiates* copying* [pnpase] Student CD
Activity 15.1 - DNA Regulatory Regions transcription factors* are needed to recognize specific DNA
sequence [motif*], binds to promoter DNA region [ activators & transcription
factors*] * makes a complimentary copy* of one of the two DNA
strands [sense strand] Quicktime movie of transcription* myDNAi Roger
Kornberg's movie of transcription (2006 Nobel)*
Kinds of RNA [table*] tRNA - small, 80n, anticodon sequence, single strand with
2ndary structure* function = picks up aa & transports it to ribosome
rRNA - 3 individual pieces of RNA - make up the organelle =
RIBOSOME primary transcript is processed into the 3 pieces of rRNA pieces (picture*) & recall structure of
ribosome
hnRNA - heterogeneous nuclear RNA : large Primary Transcript RNA function - is the precursor of mRNA in eukaryotes
hnRNA - heterogeneous nuclear RNA : large Primary Transcript RNA function - is the precursor of mRNA in eukaryotes
mRNA - intermediate sizes - 100n to 400n ( split genes*) primary transcript & mRNA
function - codes for amino acid sequence were not same size?
processing (cutting) of introns & exons*
Splicesome splicing of eucaryotic genes* [glossary] (Sumanas, Inc. advanced animation)
structure of mRNA* - caps & tails
role of 5' CAP and Poly-A Tails* [glossary] luciferase
summary of eukaryotic RNA processing*
Other classes of RNA:
small nuclear RNA (snRNP's) - plays a structural and catalytic role in spliceosome*
there are 5 snRNP's making a spliceosome [U1, U2, U4, U5, & U6];
they and participate in several RNA-RNA and RNA-protein interactions
SRP (signal recognition particle) - scRNA is a component of the protein-RNA complex
that recognizes the signal sequence of polypeptides targeted to the ER - figure*
small nucleolar RNA (snoRNA) - aids in processing of pre-rRNA transcripts for
ribosome subunit formation in the nucleolus
micro RNA's (micro-RNA) - also called antisense RNA & interfereing RNA; c7-fig 19.9*
short (20-24 nucleotide) RNAs that bind to mRNA inhibiting it. figure*
present in MODEL eukaryotic organisms as: roundworms, fruit flies, mice, humans, & plants
(arabidopsis); seems to help regulate gene expression by controlling
the timing of developmental events via mRNA action also inhibits translation of target mRNAs. ex: siRNA
--> [BARR Body*]
TRANSLATION - Making a Protein
process of making a protein in a specific amino acid sequence from a unique mRNA sequence... [ E.M.
picture* ] polypeptides are built on the ribosome (pic) on a polysome [ animation*]
Sequence of 4 Steps in Translation...
[glossary]
1. add an amino acid to tRNA -- > aa-tRNA - ACTIVATION*
2. assemble players [ribosome*, mRNA, aa-tRNA] - INITIATION*
3. adding new aa's via peptidyl transferase - ELONGATION*
4. stopping the process - TERMINATION
Review the processes - initiation, elongation, & termination myDNAi real-time movie of
translation* & Quicktime movie of translation Review figures & parts: Summary fig*
[ components, locations, AA-site, & advanced animation ]
[ Nobel Committee static animations of Central Dogma
GENETIC CODE...
...is the sequence of nucleotides in DNA, but routinely shown as a mRNA code*
...specifies sequence of amino acids to be linked into the protein
coding ratio* - # of n's... how many nucleotides specify 1 aa 1n = 4 singlets, 2n= 16 doublets, 3n = 64 triplets
Student CD Activity - 11.2 - Triplet Coding
S. Ochoa (1959 Nobel) - polynucleotide phosphorylase can make
SYNTHETIC mRNA Np-Np-Np-Np <----> Np-Np-Np + Np
Marshall Nirenberg (1968 Nobel) - synthetic mRNA's 5'-UUU-3' = phe U + C --> UUU, UUC, UCC,
CCC UCU, CUC, CCU, CUU
the Genetic CODE* - 64 triplet codons [61 = aa & 3
stop codons] universal (but some anomalies), 1 initiator
codon (AUG), redundant but non-ambiguous, and exhibits
"wobble*".
GENETIC CHANGE - a change in DNA nucleotide sequence
- 2 significant ways mutation & recombination
1. MUTATION - a permanent change in an organism's DNA*that results in
a different codon = different amino acid sequence Point mutation - a single to few nucleotides change...
- deletions, insertions, frame-shift mutations* [CAT] Student CD Activity - 11.2 -
Triplet Coding - single nucleotide base substitutions* :
non-sense = change to no amino acid (a STOP codon) UCA --> UAA ser to non
mis-sense = different amino acid UCA --> UUA ser to leu
Sickle Cell Anemia* - a mis-sense mutation... (SCA-pleiotropy)
another point mutation blood disease - thalassemia
- Effects = no effect, detrimental (lethal), +/- functionality, beneficial
2. Recombination (Recombinant DNA) newly combined DNA's that [glossary]*
can change genotype via insertion of NEW (foreign) DNA molecules into recipient cell
1. fertilization* - sperm inserted into recipient egg cell --> zygote [n + n = 2n] 2. exchange of homologous chromatids via crossing over* = new gene
combo's 3. transformation* - absorption of 'foreign' DNA by recipient cells changes
cell 4. BACTERIAL CONJUGATION* - involves DNA plasmidsg* (F+ & R =
resistance) conjugation may be a primitive sex-like reproduction in
bacteria [Hfr*] 5. VIRAL TRANSDUCTION - via a viral vector ( lysogeny* &
TRANSDUCTION* ) general transduction - pieces of bacterial DNA are
packaged w viral DNA during viral replication restricted transduction - a temperate phage goes lytic
carrying adjacent bacterial DNA into virus particle
6. DESIGNER GENES - man-made recombinant DNA molecules
Designer Genes - Genetic Engineering - Biotechnology RECOMBINANT DNA TECHNOLOGY...
a collection of experimental techniques, which allow for
isolation, copying, & insertion of new DNA sequences into
host-recipient cells by A NUMBER OF laboratory protocols & methodologies
Restriction Endonucleases-[glossary]*... diplotomic cuts* at unique DNA sequences,
Eco-R1-figure* mostly palindromes... [Never odd or even] ▼
5' GAATTC 3' 5' G . . . . . + AATTC 3' 3' CTTAAG 5' 3' CTTAA . . . . G 5'
▲ campbell 7/e movie* DNA's cut this way have sticky (complimentary) ends & can be
reannealed or spliced* w other DNA molecules to produce new genes
combos and sealed via DNA ligase. myDNAi movie of restriction
enzyme action
Procedures of Biotechnology? [Genome Biology Research]
A. Technology involved in Cloning a Gene... [animation* & the tools of genetic analysis]
making copies of gene DNA
1. via a plasmid* [ A.E. fig & human shotgun plasmid cloning & My DNAi movie]
2. Librariesg... [ library figure* & Sumanas animation - DNA fingerprint library ]
3. Probesg... [ cDNAg & reverse transcriptaseg
cDNA figure* & cDNA library* ]
4. Polymerase Chain Reactiong & figure 20.7* & animation* + Sumanas, Inc. animation*
PCR reaction protocol & Xeroxing DNA & Taq polymerase
B. Detection of a Gene... Locating a gene (or its activity) - Restriction Maps.
1. Restriction mapsg... via gel electrophoresis* &
DNA-electropherogram*
2. DNA fingerprintg... CSI Miami - how to make one* a murder case* & a rape case* + DNA prints in
Health & Society & DNA Forensic Science 3. DNA Probe Hybridizationg... to detect specific DNA
with a probe fig 20.5*
4. Comparing Restriction Fragments... to a probe Southerng Blotting fig* Sumanas, Inc. animation - DNA electrophoresis & blotting* one can detect specific gene sequence
in samples by binding to labeled probes
5. DNA micro-arrays - monitor gene expression in thousands of genes & changes
by passing cDNA of the cell's mRNA over slide with ssDNA of all cell's genes;
DNA microchips are fabricated by high speed robotics akin to Intel chip making
cDNA (mRNA's) are fluorescently tagged so easy to see in slide's wells.
[microchips arrays made simultaneously by phopshoramidite method of Caruthers]
Sumanas animation - DNA chip technology* & myDNAi DNA microarrays
5. Gene Sequencing - Human Genome Project
strategy - shotgun approach* developed by Celera Genomics
random fragments are sequenced and then ordered
relative to each other via overlap & supercomputing
Student CD Activity - 16.1 - Sequencing Strategies
methodology dideoxy procedure* (development by Fred Sanger)
Surprising Size Estimates of Human Genome & figure* NHGRI researchers* have confirmed the
existence of 19,599 protein-coding genes in the human genome and identified another
2,188 DNA segments that are predicted to be protein-coding genes =
21,787 genes
mtDNA & Y-chromosome DNA aid in search for our human ancestry
Practical Applications of DNA Technology - Some examples of What's been Do
1. Medical... disease often involves changes in gene expression a. disease/infection diagnosis:
PCR & labeled DNA probes from pathogens can help identify microbe types...
isolate HIV RNA --RT--> cDNA --PCR--> probe can id... AIDS infection
b. RFLP - Restriction Fragment Length Analysis - markers often inherited with disease
what is RFLP* genetic testing & polymorphism ---> RFLP markers to disease
DdeI cuts Sickle gene* (also MST II cuts Sickle Cell)
fragment analysis (DNA fingerprinting) also used for paternity testing
c. Gene Therapy... idea is to replace defective genes via microinjection of DNA*
requires VECTORS - fig 20.16* (patient: ADA Deficiency & Ashanti DeSilva update)
SCID (severe combined immunodeficiency - a single gene enzyme defect):
clinical trials in 2000 resulted in 2 of 9 cured, but they developed leukemia:
a retroviral vector inserted a repair gene in bone marrow cells
near genes involved in blood cell division, thus leukemia. trials stopped.
2. Pharmaceutical Products... manufactured drugs
Recombinant bacteria* = Humulin & protropin (an ethical dilemma)* Student CD Activity - 17.1
- Producing Human Growth Hormone
Control of Gene Expression How do we know a gene has been active
(turned on) within cells???? we look for gene's
product, i.e., protein or RNA
an increase in enzyme activity implies gene action?
no enzyme activity suggests no gene action
but, what about pre-existing inactive enzymes converting to --> active forms
ZYMOGENS - pepsinogen -----> pepsin
- trypsinogen -----> trypsin
thus, we have 2 possibilities: 1) pre-existing inactive
enzyme --> active 2) de novo (new) enzyme
synthesis (gene action)
Mechanism of Gene Action (turning on/off genes) in PROCARYOTES...
model: LACTOSE OPERON - Jacob & Monod
[glossary]*
E. coli (grown on) glucose lactose
NO beta-galactosidase beta-galacotsidase
OPERON* = series of mapable-linked genes controlling synthesis of protein
p Rg crp p O Sg1 Sg2 Sg3 Rg (i gene) regulator - makes repressor protein what if regulator
binds lactose* p promoter - binds RNA polymerase figure O operator - binds repressor protein figure
S structural - make enzyme proteins figure*
Sumanas, Inc. animation - Lac Operon* Catabolic Repression
Control of Gene Expression - in EUKARYOTES
Mechanism of Gene Action (turning on/off genes) is more complex
much more DNA & it's inside a compartment (nucleus) and, there are no operons present
have many more promoters - sites where RNA polymerase binds
enhancer sequence - sites where enhancers/transcription factors bind
transcription factors - proteins that help transcription but, individual genes are not contiguous, thus no operons
3 levels for eukaryotic controls* - transcriptional, translational, post-translational
multiple places for control* - of whether a gene make a protein or not
McGraw-Hill higher Ed movie on control of
gene expression
Some examples for Eukaryotic gene expression controls:
Differential Gene Activity... is the selective expression of genes
i.e., different cell types express different genes [liver vs. lens cell]
1. role of activators in selective gene expression
(Differential Gene Activity*)
ex: Steroid Hormones (figure*)
2. Molecular turnover - ½ life mRNA's* & longevity of some proteins*
3. Processing of RNA transcript (figure*) cut/spliced in nucleus and capped for
transport intron - pieces cut out (non gene-
proteins) exons - pieces transported to cytoplasm alternative splicing = figure C17.11* and
some examples*
ex. cont. Eukaryotic gene expression controls:
5. cancer often results from gene changes affecting cell cycle control.
cancer genes, such as adenomatous polyposis coli, which cause 15% of
colorectal cancers is a tumor suppressor gene, a type of Oncogenesg*
2 kinds of human cancer genes:
Ras (proto-oncogene) causes 30% human cancers: is a G-protein that promotes other cell division
proteins a Ras mutation --> hyperactive Ras protein -->
cell division fig 19.12a
p53 (tumor suppressor geneg = 50% human cancers) fig 19.12b*
p53 is a transcription factor that promotes the synthesis of cell cycle
inhibiting proteins [DNA damage --> active p53 --> p51 gene --> protein binds to
cyclin dependent kinase stops cell division]
thus a p53 mutation --> leads to excess cell division (cancer)
- other cancer genes can lead to new gene actions resulting in
cancer BRCA1 and BRCA2 (tumor suppressor genes) are involved in
50% of breast cancers in humans
Organization of the Genome* - the structural organization of genome
in eukaryotes influence its expression. Size of Human genome:
3million+ base pairs, or some 500,000 pages of journal Nature.
reading a 5 bases/sec it would take you about 60 year @ 8h/d 7d/w
A definition of a GENE*.
Definition of a Gene Mendel's Particles... unit of heredity responsible for phenotype
Morgan's Loci... he placed genes on a chromosome, i.e., it's a cellular entity, that is part of chromosome & is mapable Watson & Crick... it's a sequence of specific nucleotides along the
length of a double helical DNA molecule Molecular Definition...
length: 1 nucleotide = 0.34nm thus tRNA = 81n x 0.34 = 27.5nm
mass: 1 nucleotide = 340amu thus tRNA = 81n x 340 = 27,540amu
Modern functional definition... a DNA sequence coding for a specific
polypeptide: but, also must include... Split Genes... presence of Introns &
Exons : eukaryotic genes contain
non-coding segments (introns) and coding segments (exons
- that make proteins)
Others DNA pieces... any definition must also include: segments that code for rRNA, tRNA, & snRNP's also promoters, enhancer segments, regulator
genes, operators ? BEST ≈ "a GENE is a region of DNA that CODES for an RNA" end.
MST II restriction cuts of normal sickle beta-gene
( pink is DNA sequence & blue = 4 gel fragments) _________|__________________|CCTNAGG GAA
_____________|____________
a b c d
In 1978, Yuet Wai Kan and Andrees Dozy of the University of California-San Francisco showed that the restriction enzyme Mst II,
which cuts normal b globin DNA at a particular site, but will not recognize and therefore will not cut DNA that contains the sickle cell mutation. Mst II recognized the sequence CCTNAGG (where N = any
nucleotide). Sickle cell disease is due to a single point mutation in the beta globin gene on chromosme 11 that changes CCTGAGG to
CCTGTGG. MST II restriction cuts of recessive sickle beta-gene (blue = 3 gel
fragments) _________|___________________CCTGTGG ________________|
____________
¥ Sickle Cell disease occurs when the DNA sequence for
glutamic acid is converted to valine. This results from a change in the nucleotide T to A. This change eliminates a
site recognized by the restriction enzyme DdeI.
Restriction enzyme: DdeI (recognition sequence: 5'-C^TNAG-3')
Southern blotting probe: fragment of ß-globin coding sequence
Pattern result: normal cell = 3 fragments (1 large, a 201bp piece, and a 175bp piece
sickle cell = 2 fragments (1 large, and a 376bp piece)
fig 20.9* Thus the number of RFLP piece can indicate presence of defective alleles.
reading frame (1 codon) = CAT point mutations at hot spots - [fig]
C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-
1st point insertion or deletion
C-A-T-X-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A = mutant
2nd insertion or deletion C-A-T-X-Y-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-
C-A-T-C-A-T-C = mutant3rd insertion or deletion
C-A-T-X-Y-Z-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T-C-A-T = norma
DO NOT study the material below Gene expressions in pharmacogenomics &
toxicogenomics via microarrays 1 cM = about 1 Mb
TRANSPOSONS - pieces of DNA prone to moving & creating repeat sequences
LINE - long interspersed nuclear element holds promoter & 2 genes: RT & integrase
an anomaly - RNA Recoding*
Simple Tandem Repeats (short- 5n to 6n) or trinucleotide (3n) repeats can undergo an increase in copy
number by a process of dynamic mutation; # of tandem repeats is unique to a genetic indiv.
Variation in the length of these repeats is polymorphic. figure* individual A has ACA repeated 65 times @ loci 121, 118, and 129
individual B has a different repeat pattern at these loci STR'sa can cause genetic diseases as well:
CCG trinucleotide occur in fragile sites on human chromosomes (folate-sensitive group).
fragile X (FRAXA) is responsible for familial mental retardation. another FRAXE is responsible for a rarer mild form of mental
retardation. mutations of AGC repeats give rise to a number of neurological
disorders
3. Forensics - DNA fingerprinting is the vogue judicial modus operandi
a murder case* & a rape case* + DNA prints in Health & Society & DNA Forensic Science
DNA fingerprinting usually looks a 5 RFLP markers and blood is tested via
Southern Blotting (20.10) using probes for these alleles
4. Environmental Clean-up... bacteria can extract heavy metals (Cu, Pb,
Ni) from the environment & convert them into non-toxic compounds
genetically modified bacteria may be the "miner's" of the future
5. Franken Food... genetically modified (GM) animals & agricultural crops
Transgenics - organisms with inserted foreign DNA in their genomes
Animals* - GFP novelties* + Dolly - animal cloning companies --->
mammalian cloning success? - "pharm" animals (20.18*) --->
transgenic animal movie sheep carry human blood
protein gene that inhibits enzymes
fibrosis; artificially insemination, microinjection
of human gene, fertilized ova are put into a surrogate sheep:
chimeras mated to produce homozygote- Milk tested for active protein.
Plants - genetically modified crop plants - fig 20.19* - to get Ti plasmids in = a DNA gun* Purdue
University Gene Gun movie - Frankenfood & Edible Vaccines - National Plant Genome Initiative
Plan update future
6. Synthetic Biology... artificially manufactured biological systems
- virus models* (synthetic Biology)
ð An overview of biotechnology History of Biotechnology Human Genome Project &
Biotech Companies HHMI funded DNA Interactive
tutorial
What are Introns? and What is the Role of Intron DNA?
don't really know, but Percentage of non-coding DNA during evolution* goes up.
INTRONS - DNA Junk or sophisticated Genetic Control Elements?
Current dogma of Molecular Biology
DNA --> RNA --> Proteins, (proteins supposedly regulate gene expression) figure*
in 1977 Phillip Sharp & Richard Roberts discovered DNA contains
introns intervening DNA segments that do NOT code for proteins
a primary RNA transcript is processed by splicing to assemble protein coding exons
Presence of Introns: Absent in prokaryotes: they have few non-coding DNA sequences
as eukaryotic complexity grows so does non-coding DNA [figure]
makes up greater than 95% of the DNA less than 1.5% of human genome encodes proteins, but all of
DNA is transcribed 40% of human genome is Transposons & repeat genetic
elements.
Evolutionary Origins? may have been self-splicing mobile genetic elements
that inserted themselves into host genomes Advent of Spliceosomes: catalytic RNA/protein
complexes that snip RNAs out of mRNAs,
would encourage introns to proliferate, mutate, evolve
that inserted themselves into host genomes
Advent of Spliceosomes: catalytic RNA/protein complexes
that snip RNAs out of mRNAs,
would encourage introns to proliferate, mutate, evolve
Role of Introns? Not Junk, but rather Genetic Control Elements [figure*]
Micro RNAs - derived from introns? - occur in plants, animals, & fungi
a) help control timing of developmental processes as cell proliferation,
apoptosis, and stem cell maintenance b) help tag chromatin with methyl and acetyl
groups c) may help in alternative splicing mechanisms
COMPLEXITY: to build a complex structure one must have bricks & mortar,
as well as an architectural plan. DNA, therefore should contain both - the materials and the plan:
a) component molecules - proteins, carbs, lipids, and nucleic acids:
all known living organism use the same bricks and mortar
b) the difference between Man & Monkey is the architectural plan
Where is the Architectural Information? we've always assumed in the regulatory proteins
Maybe it's in the non-coding mirco-RNAs (intronic elements)
Thus the greater proportion of the genome of complex organisms, the introns, isn't junk,
but rather, it is functional RNA that regulates time dependent complexity?