L14: Transcriptional Regulation in Eukaryotes Flashcards
transcriptional regulation in eukaryotes vs prokaryotes
- eukaryotes have nucleosomes (comprised of histones)
- eukaryotic genes have more regulatory sequences and more transcriptional regulators
transcriptional regulation in eukaryotes vs prokaryotes - nucleosomes
- the genes are partially concealed from being expressed
- this creates an obstacle but also adds a new level of gene regulation
transcriptional regulation in eukaryotes vs prokaryotes: regulatory sequences and transcriptional regulators - promoter and regulator binding sites
- promoter (in prokaryotes): region to which transcriptional machinery binds
- regulator binding sites (in eukaryotes): DNA recognition sites bound by transcription factors
transcriptional regulation in eukaryotes vs prokaryotes - what makes regulatory binding sites so different than promoters?
- they are more in number and can be positioned at a large distance from the gene
- it allows greater integration of signals for a gene
- it is bound by activators and repressors
activator proteins - yeast transcriptional activator Gal4
- Gal4 dimerizes and activates galactose metabolism genes (GAL4)
- it binds to a regulatory sequence called UAS (upstream activating sequence)
- consists of 2 modular and separable domains
activator proteins: Gal4 - what are the 2 modular and separable domains?
- DNA-binding domain: recognizes UAS
- Activation domain: recruits transcriptional machinery
DNA-binding domains - eukaryotic transcriptional regulators
- often inserts an alpha-helix into the major groove of DNA
- termed recognition helix
DNA-binding domains example
- there are many classes
- Ex: Zinc-finger domain
DNA-binding domain example - Zinc-finger domain
- incorporates zinc atoms into structure
- alpha-helix inserted into the major groove
how can activators initiate transcription?
- recruitment of RNA pol II indirectly (prokaryotes do so directly)
- recruitment of nucleosome modifiers
how can activators initiate transcription - RNA Pol II
- interaction is through complexes such as Mediator or TFII subunits
- mediator bridges activators with RNA Pol II
how can activators initiate transcription - nucleosome modifiers
- chemically modify histones (Histone acetyltransferases of HATs) where histone acetylation loosens the chromatin to reveal DNA-binding sites
- displace or “remodel” nucleosomes (chromatin remodeling complex)
what are enhancers?
- cis-regulatory elements bound by regulatory transcription factors
- they dictate differential gene expression
- only cells that have the regulatory protein will express the gene
what are the characteristics of enhancers
- they can be 100,000 bases from the promoter and can be found in introns upstream or downstream of genes
- most genes have more than one enhancer
- enhancers can still function if moved or flipped in 5’ to 3’ orientation
explain the control of transcriptional regulator activity
- a signal input can alter the the activity of transcriptional regulators
- ex: unmasking of an activating region
control of transcriptional regulator activity - unmasking of an activating region
- Gal4 activates the GAL1 gene only in the presence of galactose (and in the absence of glucose)
- this is because galactose triggers the release of the masking protein Gal80 from Gal4
how are activators in eukaryotes different compared to prokaryotes
- multiple activators often work together synergistically by cooperative binding
- the DNA binding of one protein aids or requires the binding of a second
activator mechanisms in eukaryotes
- direct interaction
- indirectly through a common third protein
- indirectly through recruitment of a nucleosome remodeler
activator mechanisms in eukaryotes - directly
two activators (A and B) move to binding site and touch each other
activator mechanisms in eukaryotes - indirectly via a third protein
two activators (A and B) move to binding site but they can only interact with each other if protein X gets in between them
activator mechanisms in eukaryotes - indirectly via nucleosome remodeler
- one molecule (A) is bound properly but B is not due to histone wrapping DNA too tightly
- A then recruits a modifier than then causes unwinding so B can bind
why is the regulation of activators important for signal integration?
- the gene is only activated when two signals are received
- each signal is communicated by a separate activator
signal integration at HO locus in yeast
- the HO gene is only expressed in the mother cell at G1-S transition of the cell cycle
- it is regulated by 2 factors: SWI5 and SBF
signal integration at HO locus in yeast - SWI5
- it is only active in the mother cell and communicates its identity
- the binding site is too distant to activate HO expression
- it recruits chromatin modifiers that act on the SBF binding site
signal integration at HO locus in yeast - SBF
- it is only present in the bud
- it communicates the timing of G1-S transition
- it requires chromatin remodeling in order to bind near HO
- SBF binds on DNA and recruits Mediator to activate HO expression
how are repressors different in eukaryotes vs prokaryotes
eukaryotic repressors do not bind to DNA sites overlapping the promoter to block RNA Pol binding
repressor mechanisms
- competition
- inhibition
- indirect repression
repressor mechanisms - competition
the repressor binds to the region that overlaps with an activator binding site
repressor mechanisms - inhibition
the repressor binds near the activator and shields its activating region
repressor mechanisms - indirect repression
the repressor recruits a histone modifier that alters nucleosomes to inhibit transcription
indirect repression example
- GAL1 gene
- its involved in galactose metabolism (activated by Gal4)
- the factor Mig1 binds upstream of Gal1 in the presence of glucose
- Mig1 recruits Tup1 repression complex (contains HDAC)
GAL1 gene - what happens when only glucose is present?
- bc glucose is present: the Tup1 repression complex is bound to DNA
- bc galactose is not present: Gal4 is repressed by Gal80 bc it sits on top of its activation domain
- results in no GAL1 expression
GAL1 gene - what happens when galactose and glucose is present?
- bc galactose is present: Gal80 unbinds and allows the activation domain of Gal4 to be exposed
- bc glucose is present: Tup1 repression complex is bound to DNA
- results in no expression of GAL1
GAL1 gene - what happens when only galactose is present?
- the Tup1 repression one complex is not bound to DNA
- Gal80 disassociates from Gal4 allowing the activation domain to be accessible
- results in GAL1 expression
how is the GAL1 gene in eukaryotes similar to the Lac operon in prokaryotes?
two carbohydrate inputs control gene expression
