Control of Gene Expression II Flashcards
how do you get different forms of proteins from the same gene?
alternative splicing
75% of genes in humans undergo alternative RNA processing
how can you regulate RNA splicing?
via two things:
- RNA splicing can be regulated negatively by repressor molecule that prevents splicing machinery access to splice site
- RNA can be regulated positively by activating molecule that recruits and helps direct splicing machinery
speak on regulation by RNA stability
- mRNAs have a poly-A tail - confers stability
- gradual shortening of poly-A tail by exonuclease
- shortening of poly-A tail acts as a timer
- once reduced to 25 nucleotides, two pathways converge to degrade mRNA
what are the two pathways that converge to degrade mRNA?
1 - decapping: exposed mRNA degraded from 5’ end (5’ cap serves to protect RNA from RNA degrading enzymes)
2 - mRNA degraded from 3’ end through poly-A tail and into coding region
what does ferritin mRNA code for?
ferritin; used for storage of iron
what does Tfr mRNA code for?
transferrin receptor - iron absorbance (this is how cell transport iron into cells)
mRNA regulation: what happens during iron starvation?
cells do not need to store iron
so they decrease ferritin mRNA (which encodes storage proteins)
- cells must transport iron into cells; so they make more transferrin receptor (TfR) mRNA
What happens during period of iron excess?
- needs to store excess iron so the cell makes more ferritin mRNA (which makes storage proteins)
- and needs to transport LESS iron into cell; thus, less TfR mRNA is made
what governs the mRNA regulation?
IRE’s: iron responsive elements - recognition sites on mRNA for binding
and
IRPs: iron responsive regulatory protein - aconitase
what happens when the IRP binds to the IRE at 5’ ferritin mRNA?
No ferritin is translated; translation is blocked
occurs during iron starvation (when you don’t need to STORE iron, but you do need to TRANSFER it);
what happens when the IRP binds to IRE at 3’ transferrin receptor mRNA?
transferrin receptor is made - mRNA stable
occurs during iron starvation because you need iron to be TRANSFERRED in.
what happens if IRP (aconitase) does NOT bind to IRE at 5’ ferritin mRNA?
mRNA is made - you get ferritin production
essentially, aconitase binds to 5’ end in times of iron deficiency because in binding there, it stops the production of ferritin. ferritin stores the iron, and we need to USE it. not Store it. so ferritin production needs to stop, which occurs with that binding, and production of TRANSFERRITIN needs to occur - that happens when aconitase is bound to the 3’ end.
Iron causes dissociation of aconitase. so that explains why these things happen when iron is not around, and aconitase is allowed to stay bound to genes
what happens if IRP does not bind to IRE at 3’ transferrin receptor mRNA?
RNA degrades and no transferrin receptor is made; again, this happens because iron dissociates the binding of aconitase to the site - which means there’s excess iron around. we only need transferrin during times of shortage.
basically, when you have iron starvation what do you need? what about iron excess?
need more iron, need transferrin receptor, DO NOT need ferritin (storage protein).
don’t need iron, so you need to store it
which means you need ferritin
but not TfR (transferrin receptor)
whats regulation by small non-coding RNAs?
via microRNAs
they’re regulatory RNAs that regulate messenger RNA
they’re noncoding RNA; 22 nucleotides long
they silence expression of specific mRNA targets
they being to the complementary sequence in the 3’ UT end of mRNA
**degrade RNA or BLOCK TRANSLATION
overall, what does microRNA do?
stop expression of RNA
whats a key characteristic of miRNA?
one miRNA can repress hundreds of mRNAs
how can we use miRNAs to identify disease?
miRNAs change their expression profile in disease states
- eg: certain miRNAs can be elevated in stroke or cardiovascular disease
- circulating levels of miRNAs can be used to identify cancers
**miRNAs can serve as biomarkers
are the changes in microRNA expression causative of disease? or responsive to disease?
both
causative: miRNAs likely have mutations that cause disease
responsive: increased miRNA expression down regulates genes in response to disease to limit severity
whats an example of a causative miRNA expression?
tourette’s syndrome
neurological disorder manifested by motor and vocal tics
variant of SLITRK1 gene shown associated with TS
Change in recognition sequence on target SLITRK1 mRNA lead to increased miRNA binding
miR-189 binds more efficiently to target sequence in 3/UT of SLITRK1 gene and decreases SLITRK1 expression- leading to TS
List a few post-translational processes and modifications that proteins go through
- modified by protein kinases
- glycosylated
- bind to other protein subunits or protein partners
- modifying enzymes act on proteins
what happens when theres an increase in temperature?
proteins misfold; so heat shock proteins are needed
these molecular chaperones are synthesized when temps rise because as temps rise, proteins misfold. Hsp’s help proteins refold.
two types: hsp60 and 70
how else do you regulate proteins?
garbage disposal via the proteasome
proteasomes can control protein activity by choosing what proteins are around
they remove misfolded proteins
how are proteins recognized by the proteasome?
ubiquitin
its a recognition tag that leads to the removal of abnormal or misfiled proteins
how does ubiquitin get placed on protein?
E1 ubiquitin activating enzyme is involved in the first step
linkage of ubiquitin to CYSTEINE side chain on E1 enzyme
then ubiquitin is transferred to E2 ubiquitin conjugating enzyme (with E3 ubiquitin ligase)
- the complex is now primed to mark proteins for destruction
how does the addition of protein to ubiquitin ligase occur?
ubiquitin chain added to LYSINE side chain on protein
each successive ubiquitin added to lysine of ubiquitin chain starts with E1 enzymes
- the targeted ubiquitin chain on target protein is recognized by proteasome
what is a proteasome?
an apparatus that deliberately destroys aberrant proteins
can proteasome be used for therapy?
yes
what does it help treat? how?
multiple myelomas
simply put: by inhibiting the activity of proteasomes on pro-apoptotic factors, which allows for apoptosis to occur
what is meyloma?
cancer of plasma cells
what is the inhibitor of proteasome called?
bortezomib - highly effective
how does it work?
chamber in proteasome has 3 proteolytic sites
bortezomid interacts with 1 proteolytic sit - reversibly inhibiting the proteasome
this inhibition leads to the prevention of degradation of pro-apoptotic factors for cell suicide = so it triggers programmed cell death in neoplastic cancer cells
specifically targets myeloma cells
how do you activate a ubiquitin ligase?
1) phosphorylation by protein kinase
2) allosteric transition cased by ligand binding
3) allosteric transition caused by protein subunit addition
how do you activate a target protein degradation signal?
1) phosphorylation via protein kinase
2) unmasking by protein dissociation
3) creation of destabilizing N-terminus
what are 4 other controls of gene expression?
1) coordinated expression of genes: genes do not exist in a vacuum
2) decisions for specialization: what kind of cell do i want to become
3) methylation and genomic imprinting: what genes get expressed (or not) from mom and dad
4) X- chromosome inactivation
coordinated gene expression - how does it occur/
expression of critical regulatory protein can trigger battery of downstream genes
- coordinated expression in response TO NEED
- eg: glucocorticoid cortisol - it responds to high stress - glucocorticoid hormone - by amplifying gene expression
decision for specialization
combinations of gene control can produce many types of genes
eg: hematopoiesis
HSC: hematopoietic stem cell; from here, you get RBC and all WBCs
DNA methylation, what does it do?
represses gene expression via covalent bond
speak on the inheritance of DNA methylation
methylation of cytosine occurs at CG sequences
methylation is inherited - cytosine is methylated by “maintenance methyltransferase”
- DNA methylation of parent strand serves as template for daughter strand
what is genomic imprinting based on?
dna methylation
what is genomic imprinting?
differential expression of genetic material depending on the parent of origin
what is epigenetic?
the regulation of expression of gene activity without altering gene structure (eg methylation)
what is one type of genomic imprinting disorder? elaborate on it
Prader Willi syndrome (PWS): caused by paternal deletion on chromosome 15
subjects inherit gene deletion from father; has unusual presentation
stage one: infantile hypotonia; poor suck; feeding difficulties - failure to thrive
stage two: hyperphagia (uncontrollable eating); onset of early childhood obesity
How does genomic imprinting play a role in PWS?
paternal gene expression: when genes in this chromosomal region are not expressed when inherited from mom but expressed when inherited from dad
normal individual: gene is expressed
PWS: paternal gene not expressed (deleted) and maternal gene not expressed even though they are present
- result of paternal deletion is lack of gene expression due to genomic imprinting
x chromosome inactivation
dosage compensation occurs so equal number of genes expressed from X chromosome is equal in males and females
when one X chromosome is inactivated in females
what is a Barr body?
when the X chromosome becomes condensed, at random.
so, heterochromatin
