lysogenic vs lytic life cycle 2007-2008 prokaryotic (bacterial) genes
TRANSCRIPT
Bacterial metabolism• Bacteria need to respond quickly to changes in
their environment– if they have enough of a product,
need to stop production• why? waste of energy to produce more• how? stop production of enzymes for synthesis
– if they find new food/energy source, need to utilize it quickly• why? metabolism, growth, reproduction• how? start production of enzymes for digestion
STOP
GO
Different way to Regulate Metabolism• Gene regulation – instead of blocking
enzyme function, block transcription of genes for all enzymes in tryptophan pathway• saves energy by
not wasting it on unnecessary protein synthesis
= inhibition-
Now, that’s a good idea from a lowly bacterium!
--
Gene regulation in bacteria• Cells vary amount of specific enzymes by
regulating gene transcription– turn genes on or turn genes off• turn genes OFF example
if bacterium has enough tryptophan then it doesn’t need to make enzymes used to build tryptophan• turn genes ON example
if bacterium encounters new sugar (energy source), like lactose, then it needs to start making enzymes used to digest lactose
STOP
GO
Bacteria group genes together • Operon – genes grouped together with related functions
• example: all enzymes in a metabolic pathway– promoter = RNA polymerase binding site
• single promoter controls transcription of all genes in operon• transcribed as one unit & a single mRNA is made
– operator = DNA binding site of repressor protein
So how can these genes be turned off?• Repressor protein– binds to DNA at operator site – blocking RNA polymerase– blocks transcription
So how can these genes be turned off?• Repressor protein– binds to DNA at operator site – blocking RNA polymerase– blocks transcription
operatorpromoter
Operon model
DNATATA
RNApolymerase
repressor
repressor = repressor protein
Operon: operator, promoter & genes they controlserve as a model for gene regulation
gene1 gene2 gene3 gene4RNApolymerase
Repressor protein turns off gene by blocking RNA polymerase binding site.
1 2 3 4mRNA
enzyme1 enzyme2 enzyme3 enzyme4
mRNA
enzyme1 enzyme2 enzyme3 enzyme4operatorpromoter
Repressible operon: tryptophan
DNATATA
RNApolymerase
tryptophan
repressor repressor protein
repressortryptophan – repressor proteincomplex
Synthesis pathway modelWhen excess tryptophan is present, it binds to tryp repressor protein & triggers repressor to bind to DNA– blocks (represses) transcription
gene1 gene2 gene3 gene4
conformational change in repressor protein!
1 2 3 4
repressortrpRNApolymerase
trp
trp
trp trp
trp trp
trptrp
trptrp
trp
Tryptophan operonWhat happens when tryptophan is present?Don’t need to make tryptophan-building enzymes
Tryptophan is allosteric regulator of repressor protein
mRNA
enzyme1 enzyme2 enzyme3 enzyme4operatorpromoter
Inducible operon: lactose
DNATATARNApolymerase
repressor repressor protein
repressorlactose – repressor proteincomplex
lactose
lac repressor gene1 gene2 gene3 gene4
Digestive pathway model When lactose is present, binds to lac repressor protein & triggers repressor to release DNA– induces transcription
RNApolymerase
1 2 3 4
lac lac
laclac
laclac
lac
conformational change in repressor protein!
lac
lac
Lactose operonWhat happens when lactose is present?Need to make lactose-digesting enzymes
Lactose is allosteric regulator of repressor protein
Jacob & Monod: lac Operon• Francois Jacob & Jacques Monod– first to describe operon system– coined the phrase “operon”
1961 | 1965
Francois JacobJacques Monod
Operon summary• Repressible operon – usually functions in anabolic pathways
• synthesizing end products
– when end product is present in excess,cell allocates resources to other uses
• Inducible operon – usually functions in catabolic pathways,
• digesting nutrients to simpler molecules
– produce enzymes only when nutrient is available• cell avoids making proteins that have nothing to do, cell
allocates resources to other uses
Positive gene control
• occurs when an activator molecule interacts directly with the genome to switch transcription on.
• Even if the lac operon is turned on by the presence of allolactose, the degree of transcription depends on the concentrations of other substrates.
• The cellular metabolism is biased toward the utilization of glucose.
Positive Gene Regulation
• Some operons are also subject to positive control through a stimulatory protein, such as catabolite activator protein (CAP), an activator of transcription
• When glucose (a preferred food source of E. coli) is scarce, CAP is activated by binding with cyclic AMP
• Activated CAP attaches to the promoter of the lac operon and increases the affinity of RNA polymerase, thus accelerating transcription
Positive Gene Regulation– If glucose levels are
low (along with overall energy levels), then cyclic AMP (cAMP) binds to cAMP receptor protein (CRP) which activates transcription.
• If glucose levels are sufficient and cAMP levels are low (lots of ATP), then the CRP protein has an inactive shape and cannot bind upstream of the lac promotor.
The BIG Questions…• How are genes turned on & off
in eukaryotes?• How do cells with the same genes
differentiate to perform completely different, specialized functions?
Evolution of gene regulation• Prokaryotes– single-celled– evolved to grow & divide rapidly– must respond quickly to changes in external
environment• exploit transient resources
• Gene regulation– turn genes on & off rapidly• flexibility & reversibility
– adjust levels of enzymes for synthesis & digestion
Evolution of gene regulation• Eukaryotes– multicellular– evolved to maintain constant internal
conditions while facing changing external conditions• homeostasis
– regulate body as a whole• growth & development
– long term processes• specialization
– turn on & off large number of genes• must coordinate the body as a whole rather than
serve the needs of individual cells
Points of control• The control of gene expression
can occur at any step in the pathway from gene to functional protein1. packing/unpacking DNA
2. transcription
3. mRNA processing
4. mRNA transport
5. translation
6. protein processing
7. protein degradation
1. DNA packing as gene control• Degree of packing of DNA regulates transcription– tightly wrapped around histones
• no transcription• genes turned off heterochromatin
darker DNA (H) = tightly packed euchromatin
lighter DNA (E) = loosely packed
H E
DNA methylation• Methylation of DNA blocks transcription factors – no transcription
genes turned off– attachment of methyl groups (–CH3) to cytosine
• C = cytosine– nearly permanent inactivation of genes
• ex. inactivated mammalian X chromosome = Barr body
Histone acetylation Acetylation of histones unwinds DNA
loosely wrapped around histones enables transcription genes turned on
attachment of acetyl groups (–COCH3) to histones
conformational change in histone proteins transcription factors have easier access to genes
Epigenetic Inheritance
• Although the chromatin modifications just discussed do not alter DNA sequence, they may be passed to future generations of cells
• The inheritance of traits transmitted by mechanisms not directly involving the nucleotide sequence is called epigenetic inheritance
2. Transcription initiation
• Control regions on DNA– promoter• nearby control sequence on DNA• binding of RNA polymerase & transcription factors• “base” rate of transcription
– enhancer• distant control
sequences on DNA• binding of activator
proteins• “enhanced” rate (high level)
of transcription
Model for Enhancer action
• Enhancer DNA sequences – distant control sequences
• Activator proteins – bind to enhancer sequence &
stimulates transcription
• Silencer proteins – bind to enhancer sequence & block
gene transcription
Turning on Gene movie
Transcription complex
Enhancer
ActivatorActivator
Activator
Coactivator
RNA polymerase II
A
B F E
HTFIID
Core promoterand initiation complex
Activator Proteins• regulatory proteins bind to DNA at distant
enhancer sites• increase the rate of transcription
Coding region
T A T A
Enhancer Sitesregulatory sites on DNA distant from gene
Initiation Complex at Promoter Site binding site of RNA polymerase
Fig. 18-9-3
Enhancer TATAbox
PromoterActivators
DNAGene
Distal controlelement
Group ofmediator proteins
DNA-bendingprotein
Generaltranscriptionfactors
RNApolymerase II
RNApolymerase II
Transcriptioninitiation complex RNA synthesis
3. Post-transcriptional control• Alternative RNA splicing– variable processing of exons creates a family of
proteins
4. Regulation of mRNA degradation• Life span of mRNA determines amount of
protein synthesis– mRNA can last from hours to weeks
RNA processing movie
5. Control of translation• Block initiation of translation stage – regulatory proteins attach to 5' end of mRNA • prevent attachment of ribosomal subunits & initiator
tRNA• block translation of mRNA to protein
Control of translation movie
6-7. Protein processing & degradation
• Protein processing– folding, cleaving, adding sugar groups,
targeting for transport• Protein degradation– ubiquitin tagging– proteasome degradation
Protein processing movie
Ubiquitin• “Death tag”– mark unwanted proteins with a label – 76 amino acid polypeptide, ubiquitin– labeled proteins are broken down rapidly in
"waste disposers"• proteasomes
1980s | 2004
Aaron CiechanoverIsrael
Avram HershkoIsrael
Irwin RoseUC Riverside
Proteasome • Protein-degrading “machine”– cell’s waste disposer– breaks down any proteins
into 7-9 amino acid fragments• cellular recycling
play Nobel animation
Concept 18.3: Noncoding RNAs play multiple roles in controlling gene
expression• Only a small fraction of DNA codes for proteins,
rRNA, and tRNA• A significant amount of the genome may be
transcribed into noncoding RNAs• Noncoding RNAs regulate gene expression at two
points: mRNA translation and chromatin configuration
RNA interference• Small interfering RNAs (siRNA)– short segments of RNA (21-28 bases)• bind to mRNA• create sections of double-stranded mRNA• “death” tag for mRNA– triggers degradation of mRNA
– cause gene “silencing”• post-transcriptional control• turns off gene = no protein produced
NEW!
siRNA
Action of siRNA
siRNA
double-stranded miRNA + siRNA
mRNA degradedfunctionally turns gene off
Hot…Hotnew topicin biology
mRNA for translation
breakdownenzyme(RISC)
dicerenzyme
initiation of transcription
1
mRNA splicing
2
mRNA protection3
initiation of translation
6
mRNAprocessing
5
1 & 2. transcription - DNA packing - transcription factors
3 & 4. post-transcription - mRNA processing
- splicing- 5’ cap & poly-A tail- breakdown by siRNA
5. translation - block start of translation
6 & 7. post-translation - protein processing - protein degradation
7 protein processing & degradation
4
4
Gene Regulation
Molecular Biology of Cancer• Oncogene
•cancer-causing genes• Proto-oncogene
•normal cellular genes• How?
1-movement of DNA; chromosome fragments that have rejoined incorrectly 2-amplification; increases the number of copies of proto-oncogenes
• 3-proto-oncogene point mutation; protein product more active or more resistant to degradation
• Tumor-suppressor genes •changes in genes that prevent uncontrolled cell growth (cancer growth stimulated by the absence of suppression)
Cancers result from a series of genetic changes in a cell lineage
– The incidence of cancer increases with age because multiple somatic mutations are required to produce a cancerous cell
– As in many cancers, the development of colon cancer is gradual