Chapter 9: Regulation of Transcription Flashcards
key to providing the cell with the correct amount of gene product at the correct time
transcriptional regulation
transcriptional regulation underlies () and employs a wide range of mechanisms
cell differentiation and development
one level of transcriptional regulation is (1), but most regulation comes from (2)
- promoter strength
- targeted gene regulation
targeted gene regulation can happen at (3)
- transcription initiation (most prevalent)
- elongation or termination
- regulation from transcribed RNA itself
regulatory (1) regulate transcription by binding to regulatory (2)
- proteins
- sequences
control relative amount of transcription from promoter
regulatory proteins
2 types of regulatory proteins
- repressors - decrease transcription level
- activators - increase transcription level
specific DNA regions to which regulatory proteins bind
regulatory sequences
in bacteria, sequences recognized by regulatory proteins are called ()
operator sites
bacterial operator sites are typically ()
close to or overlap promoter
if operators or (and common for enhancers) are distal to the gene, DNA must loop around for the regulatory protein to interact with the polymerase, sometimes aided by ()
architectural DNA binding proteins
eukaryotic analogs to bacterial operator sites; usual distal (1000s of bps away) from genes
enhancers
to increase the sophistication and subtlety of regulation, eukaryotic regulatory sequences frequently bind ()
several regulatory proteins
found in more complex eukaryotes; have a combination of enhancer and insulator elements
locus control regions (LCRs)
blocks unwanted interaction between enhancers and certain genes
insulators
role of LCRs in the expression of beta-globin genes
LCRs make sure that only fetal Hb is expressed while in the womb, and only adult Hb is expressed outside of the womb
each regulatory protein has a () that recognizes a specific sequence
DNA binding domain
regulatory proteins are often (), allowing them to have additional domains that give them additional functions
modular
transcription level changes in response to (1); this can be done by changing the (2) of the regulatory proteins
- changing conditions
- activity
small molecules that can bind directly to regulatory proteins and change their conformation upon binding
allosteric effectors
() can also change how regulatory proteins interact
phosphorylation or other covalent modifications
covalent modification, along with other factors such as () serve to fine-tune regulation of gene expression
regulator abundance and localization
() chromatin tends to be actively transcribed
hyperacetylated
hypoacetylated chromatin tends to have () levels of transcription
low
other histone modifications like (4) also affect transcription, but the relationship is not yet clear
- methylation
- phosphorylation
- ubiquitination
- sumoylation
a () has been proposed (and is being actively researched) in which certain combinations of modifications would lead to specific outcomes
histone code
() are tailored to fit DNA; some are found throughout life, other are found just in eukaryotes or metazoans
DNA-binding motifs
the () is a common DNA-binding motif and is found in many protein folds
helix-turn-helix
the second helix in the helix-turn-helix motif is called the (), and fits into the major groove of DNA
recognition helix
recognition helices in helix-turn-helix motifs read the DNA sequence through ()
interactions with the base pairs
many helix-turn-helix DNA-binding motifs are dimers, with the recognition helices spaces () apart to fit in neighboring grooves
3.4 nm
the () is a monomeric helix-turn-helix, commonly found in eukaryotes; its recognition helix is longer than its bacterial counterparts
homeodomain
the sidechains of homeodomain motifs interact with (1), while its N-terminal arm makes contacts in the (2)
- DNA base pairs
- minor groove
a prominent part of many DNA binding folds -> most common DNA-binding domain in the human genome; has a domain of ~30 amino acids
zinc finger
structure of a zinc finger
alpha helix and 2 beta strands around a central zinc ion
the zinc ion in zinc fingers interacts with (); these are always conserved in proteins with zinc fingers
2 cysteines and 2 histidines -> Cys2His2 zinc fingers
each zinc finger inserts its alpha helix into the (1) of DNA and recognizes (2)
- major groove
- 2-3 base pairs
how do proteins with zinc fingers enhance specificity when recognizing DNA sequences
proteins have multiple zinc fingers -> zinc fingers are multimerized
alpha helices that wind around each other; form part of some DNA-binding domains
coiled-coils
2 main examples of coiled-coil motifs in DNA-binding domains
- basic region-leucine zipper (bZIP) proteins
- basic region-helix-loop-helix (bHLH) proteins
structure of bZIP proteins
- 2 alpha helices about 60 AAs long
- helices have hydrophobic leucine residues
- helices splay and sit in the DNA major groove at the N terminal end
structure of bHLH proteins
- 4 helices joined by a loop (four-helix bundle)
both bZIP and bHLH proteins become () when not bound to DNA
unstructured
the () binds DNA with 2 beta strands
MetJ repressor
(1), part of the mammalian transcriptional regulator nuclear factor (2), are largely beta sheets
- p50 and p65
- NF-kappa(B)
() are entirely made up of beta sheets and connecting loops
immunoglobulins
a simple way to regulate transcription is to ()
prevent RNA pol from accessing the promoter
bacterial genes that are located, regulated, and transcribed together
operon
the () is a protein with a helix-turn-helix DNA-binding motif that responds to the level of tryptophan in the cell
Trp repressor
when tryptophan levels are high in the cell, Trp repressor binds to () and blocks RNA pol from binding
Trp operator
Trp repressor can only bind to DNA when (1), which only happens when (2)
- it is bound to Trp
- Trp levels are high
regulatory protein that enhances the binding of RNA pol to a promoter to increase transcription levels
activators
() activates more than 100 E. coli promoters when carbon sources are low
catabolite activator protein (CAP)
the catabolite activator protein (CAP) is also called
cAMP receptor protein (CRP)
() leads to increase in cAMP levels -> cAMP binds to CAP and increases CAP’s binding to DNA
glucose depletion
CAP enhances RNA pol binding to DNA by binding to the ()
CTD
2 classes of promoters that interact with CAP
- class I - CAP binds upstream of promoter
- class II - CAP binding site overlaps the RNA pol binding site
() responds to low glucose levels in the cell by allowing the cell to metabolize lactose instead (if lactose is present) -> contains genes that are needed for metabolizing lactose
lac operon
(1) and (2) regulate the lac operon
- CAP
- LacI repressor
how does the LacI repressor regulate the lac operon
- when lactose is present, it binds to the LacI repressor and prevents it from binding to operator
- when lactose is absent, LacI is able to bind to lac opertor to prevent transcription of lac operon
how does CAP activator regulate the lac operon
- glucose present: decreased cAMP levels -> cAMP doesn’t bind to CAP and CAP doesn’t bind to lac operator
- glucose absent: increased cAMP levels -> cAMP binds CAP and CAP binds to lac operator to increase transcription of lac operon
in () conditions, transcription for lac operon is strongly on
low glucose, available lactose
in (1) conditions, there is weak transcription of lac operon; aka (2)
- high glucose, available lactose
- basal transcription
if the spacing between -10 and -35 promoter elements is (), RNA pol binding is weakened -> another layer of regulation
very large
() can change the way promoter elements are spatially related to each other, thereby enhancing transcription
MerR family regulators
the MerR protein can bind to the DNA and change its conformation to ()
a spacing more favorable for RNA pol binding
RNA pol holoenzyme that contains sigma 54 needs to be activated by () to drive promoter opening
ATP hydrolysis
() is an example of an activator with sigma 54
NtrC
NtrC binds at enhancer elements and activates transcription when ()
phosphorylated
phosphorylation triggers () of NtrC, and promotes its interaction with the polymerase at the promoter
oligomerization
in NtrC activation, oligomerization stimulates NtrC’s (), which helps promote the formation of the open complex
rate of ATP hydrolysis
responses to external signals often involve () that leads to a signaling cascade
phosphorylation of a receptor protein
a 2-component signal transduction pathway is an example of how phosphorylation of a receptor protein leads to a ()
signaling cascade
2-component signal transduction pathway have a (1) and a (2)
- sensor kinase
- response regulator
the sensor kinase in a 2 component transduction pathway has a () that becomes autophosphorylated on receipt of the signal
histidine (histidine kinases)
in 2 component transduction pathways, the phosphoryl group on an autophosphorylated histidine is transferred to an (1) in a (2)
- aspartic acid residue
- response regulator protein
Complex and differing signals regulate transcription. The outcome of the combination of signals, which can be (1), depends on the (2)
- antagonistic
- strength of the opposing regulators.
() infects E. coli by injecting its chromosome into the
bacterium
Bacteriophage lambda
after infecting E. coli, bacteriophage lambda enters either (1) or (2)
- lytic growth
- lysogeny
when bacteriophage lambda infect E. coli, () produces lots of copies of the virus in the bacterial cell and eventually leads to cell lysis
lytic growth
lytic growth allows a phage to () a cell that may not survive
escape
when bacteriophage lambda infects E. coli, () happens when the virus integrates into the host genome and becomes dormant
lysogeny
in lysogeny, the integrated virus is called a ()
prophage
lysogeny allows bacteriophage lambda to multiply in conditions where ()
there might not be enough nearby cells to infect
when E. coli containing prophage endures (1), the phage can be triggered to excise itself and (2)
- difficult conditions (e.g. DNA damage)
- undergo lytic growth
lysis and lysogeny are regulated by levels of the DNA-binding proteins (3)
- cI (lambda repressor)
- cII
- Cro
cI, cII, and Cro bind to sites in the phage chromosome and control transcription at the ff 4 promoters:
- P_R
- P_L
- P_RE
- P_RM
(2) are the early promoters involved in the choice between lysis and lysogeny -> host RNA pol initiates basal transcription
P_R and P_L
P_R promoter encodes (1) regulators, while P_L encodes (2)
- Cro and cII
- N protein
() prevents premature termination of the P_R transcript and thereby allows transcription of Cro and cII
N protein
if sufficient cII accumulates, it stimulates transcription at (1), which produces (2)
- P_RE
- cI
cI stimulates (), which makes more cI
P_RM
high cI levels result in cI binding to P_R and P_L promoters -> repress (1) and activates transcription of (2)
- lytic genes
- integrase
() promote and maintain lysogeny
high cI levels
if not enough cII is produced, (1) binds to P_RM and represses (2)
- Cro
- cI production
by preventing (), Cro ensures the expression of lytic genes from P_R and P_L
cI synthesis
(1) regulate the levels of cII -> cII is sensitive to host cell (2)
- environmental factors
- proteases
if proteases are (), cII is degraded -> cI cannot accumulate and virus enters (2)
- highly active
- lytic growth
the () of cI and Cro at 6 DNA-binding sites determine the choice between lysogeny and lysis
binding affinities
cI has 2 domains that can form (1) and (2)
- tetramers
- oligomers
Cro has a single small domain that binds as a ()
dimer
operators O_L and O_R (for promoters P_L and P_R, respectively) each have () that cI or Cro can bind to
3 sites
cI binds first to operators (), where it has strongest affinity
OL1 and OR1
binding of cI to OL1 and OR1 recruis another cI to weaker sites ()
OL2 and OR2
the cI units bound to operators OL1-2, OR1-2 (1) to form a (2)
- oligomerize
- looped-out structure
oligomerization of the cI units at operator sites stops (1) and promotes (2) -> increase in cI levels
- RNA pol binding to PL and PR
- transcription from PRM
with increasing levels of cI, cI binds to (), stoping the activation of PRM -> suppressing cI synthesis
OL3 and OR3
binding of cI to OL3 and OR3 serves as ()
autoregulation
if there is insufficient cI, Cro accumulates and binds with the greatest affinity to (), blocking PRM and suppressing cI synthesis
OR3
more Cro synthesis leads to binding at the other (1), autoregulating Cro by preventing RNA pol binding to (2)
- OR and OL sites
- PR promoter
the prophage must be able to excise itself and switch from lysogeny to lytic growth if the host cell is ()
threatened
excision of prophage is brought about by a response to () that disables cI
DNA damage
DNA damage in a bacteriophage infected bacterium prompts a (1), activating (2)
- host SOS response
- RecA
cleaves cI so it cannot dimerize or cooperate with other cI units
RecA
after cI cleavage by RecA, cI dissociates from (), allowing transcription from PL and PR-> allowing lytic growth
operator sites
regulation can also occur at the () steps of transcript synthesis
elongation and termination
some phage lambda genes are transcribed only when transcription termination is actively prevented -> this is called ()
anti-termination
() prevent stalling and termination by altering the properties of bacterial RNA pol
bacteriophage lambda N and Q proteins
initially, transcription from PL and PR promoters successfully makes Cro and N protein until RNA pol encounters termination sites ()
tL and tR
when N protein builds up, it stops ()
termination of transcription from PL and PR promoters
N proteins bind to (1) in the transcribed RNA in a (2)
- nut sites
- step-loop form
when N protein binds to nut sites, it allows several proteins to be (), allowing RNA pol to transcribe through tL and tR
recruited and interact with RNA pol
with termination at tL and tR suppressed (due to build up of N proteins), downstream genes can be transcribed, including ()
bacteriophage lambda protein Q
() overrides termination at the PR promoter, allowing transcription of late lytic genes
Q protein
in contrast to N protein, Q prevents termination by ()
binding directly to DNA
the Q binding site (QBE) is between the () of the PR promoter
-35 and -10 elements
without Q protein, transcription terminates very early, just downstream of ()
-10
Q protein directly contacts the (1) and allows elongation to resume; Q remains associated with the (2)
- sigma subunit (sigma70)
- elongation complex
() controls bacterial genes needed for amino acid biosynthesis
attenuation
attenuation exploits ()
folding of RNA into alternative secondary structures
the trp operon is controlled by Trp levels in the cells -> different levels cause the transcript to have ()
different secondary structures
the genes for Trp synthesis are transcribed together -> a ()
polycistronic message
the mRNA for Trp synthesis has a leader sequence near the start that has an (1) and 2 sequences that can form (2)
- intrinsic terminator; an attenuator
- stem-loops
the leader sequence in mRNA for Trp synthesis has a () with lots of codons for Trp
small open reading frame
the leader sequence in mRNA for Trp synthesis has 4 blocks that can form alternative pairing arrangements: 1 + 2
stem-loop
the leader sequence in mRNA for Trp synthesis has 4 blocks that can form alternative pairing arrangements: 3 + 4
Rho-independent terminator
the leader sequence in mRNA for Trp synthesis has 4 blocks that can form alternative pairing arrangements: 2 + 3 (1 and 4 are unpaired)
stem-loop
in low Trp conditions, ribosome stalls while translating the peptide from leader sequence of mRNA (for Trp synthesis) -> region 1 is (a), region 2 and 3 (b), preventing 3 and 4 from (c), so transcription continues
a. blocked
b. form a stem-loop
c. binding and forming a terminator
() are portions of a transcript that can directly bind a small molecule that controls the RNA secondary structure -> regulates transcription or translation
riboswitches
riboswitches have 2 regions:
- aptamer - binds to metabolite
- expression platform - controls transcription or translation
the () regulates adenine synthesis and transport -> gene expression depends on whether a terminator or anti-terminator forms
B. subtilis adenine riboswitch
what happens to B. subtilis adenine riboswitch in low adenine conditions
regions 2 and 3 form anti-terminator -> transcription proceeds
what happens to B. subtilis adenine riboswitch in high adenine conditions
regions 3 and 4 form a terminator
Eukaryotes typically regulate transcription via DNA-binding proteins that recruit (1) or (2)
- co-activators
- co-repressors
example of eukaryotic DNA-binding protein that recruits co-activator/co-repressor
ELK1, which recruits Mediator
extracellular signals that promote entry into mitosis
mitogens
in the absence of mitogens, ELK1 binds to () but doesn’t activate transcription
serum response factor (SRF)
Mitogen binding at the cell surface activates kinases, which ()
phosphorylate ELK1
phosphorylated ELK1 recruits (), promoting transcription
mediator
() regulates genes that are responsible for galactose metabolism
Gal4
Gal4 dimer activates transcription by binding to the ()
UASG sequence
Gal4 activity is regulated by (), which respond to galactose in the cell
Gal80 and Gal3
Unlike bacterial Trp repressor and CAP activator, the effector (galactose) for Gal80 and Gal3 doesn’t ()
bind directly to Gal4.
in the absence of galactose, () binds to activating domain of Gal4, preventing transcription
Gal80
in the absence of galactose, Gal80 is found in (1), while Gal3 is found in (2)
- nucleus and cytoplasm
- cytoplasm only
in presence of galactose, galactose binds to (1), which allows (1) to bind to (2)
- Gal3
- Gal80
in presence of galactose, Gal3 sequesters Gal80 in the ()
cytoplasm
with Gal80 sequestered in the cytoplasm, Gal4 is able to recruit () and Mediator, which activates transcription
SAGA (co-activator complex)
() responds to nutritional cues in yeast and can activate or repress transcription
Ume6
where there is enough (1) in the cell, Ume6 binds DNA and recruits co-repressors (2)
- Nitrogen and Carbon
- Sin3, Rpd3, Isw2
(): a histone deacetylase – histone deacetylation promotes more compact chromatin, which represses transcription
Rpd3
(): a nucleosome remodeling enzyme, which helps establish the altered chromatin pattern
Isw2
Ume6 is (1) in the absence of N and C, (2) dissociate
- phosphorylated
- Sin3 and Rpd3
additionally, in the absence of N and C, a co-activator, () is recruited
Ime1
() is involved in protecting the cell during abnormally high temperatures (heat shock) -> particularly observed in Drosophila
Hsp70
promoter proximal pausing is used to prime Hsp70 (heat shock protein 70) for rapid transcription in response to ()
heat shock
without heat shock at 25C, () binds upstream of hsp70
GAGA factor
without heat shock at 25C, GAGA factor recruits (), which keeps the promoter free of nucleosomes and allows polymerase binding
NURF
But RNA Pol is paused by negative elongation factors () -> polymerase complex is not phosphorylated enough
NELF, DSIF
in transcription of Hsp70, without heat shock, Hsf monomers are present in the cell but ()
cannot bind to DNA
with heat shock at 37C, Heat-Shock Factor (Hsf) trimerizes and binds to ()
heat shock elements (HSEs)
when bound to HSEs, Hsf recruits Mediator and ()
p-TEFb (kinase)
recruitment of Mediator and p-TEFb by Hsf () Pol Rpb1, NELF, DSIF -> resumes elongation
phosphorylates
() also depends on anti-termination (similar to phage lambda N protein)
HIV transcription
in HIV transcription, the (1) forms a step-loop called the (2) -> leads to transcription termination
- tar site
- TAR element
in HIV transcription, transcription termination occurs if () is absent
Tat protein
() binds to tar site in HIV transcription along with a cellular kinase -> phosphoryltates RNA pol II CTD -> relieves pausing and precents premature transcript termination
Tat (viral protein)
- regulation of a single gene with several different proteins, each of which is controlled by a different parameter
combinatorial control
combinatorial control is the response of a gene to ()
multiple signals and regulatory proteins
yeast cell type is controlled by () transcriptional factors
four
yeast cell has 3 cell types:
- a
- alpha
- a/alpha
() yeast cell types are haploid
a and alpha
(1) type yeast cells are diploid and result from mating of (2)
- a/alpha
- a and alpha
a/alpha yeast cells cannot mate, but can () when starved
undergo meiosis
a. alpha, and a/alpha yeast cells have distinctive gene expression patterns, regulated by 4 proteins:
a1, alpha1, alpha2, and MCM1
regulatory proteins for yeast cells: repressors
a1 and alpha2
regulatory proteins for yeast cells: activators
alpha1, MCM1
in yeast cells, a1, alpha1, and alpha2 proteins are encoded at the (1) on (2)
- MAT locus
- chromosome III
in yeast cells, the MAT loci are different in different cell types:
1. a cells encode: (a)
2. alpha cells encode (b)
3. all 3 types encode (c)
a. a1
b. alpha1/2
c. MCM1
in a-type yeast cells, MCM1 activates (1), while (2) are off
- a-specific genes
- alpha-specific
in alpha-type yeast cells, (1) activates alpha-specific genes; (2) represses a-specific genes
- alpha1-MCM1
- alpha2-MCM1
in alpha yeast cells, MCM1 activates transcription when bound by (1), and represses transcription when bound to (2)
- alpha1
- alpha2
in a/alpha yeast cells, (1) form a heterodimer and repress haploid-specific genes: (2)
- a1-alpha2
- alpha1 and RME1
() represses meiosis genes in yeasts; if this is no longer expressed, meiosis can occur
RME1
Several proteins regulate (). This is important in viral defense by inhibiting synthesis of viral genome and stimulating host immune response)
human interferon-β
A number of proteins bind to the interferon-β enhancer to form an ()
enhanceosome
() between neighboring proteins are required for stable complex assembly involving at the interferon-β enhancer
Cooperative interactions
binding of proteins at the interferon-β enhancer are aided by an architectural DNA-binding protein – () -» induces DNA bending, which helps other factors bind
HMG-I(Y)
Once the enhanceosome is complete, the interferon-β gene is activated -> requirement of using a large number of proteins to activate a single gene ensures that the gene is activated only under ().
a very precise set of conditions
extracellular signals trigger biochemical events whose end result is a change in gene expression
signaling cascades
() respond specifically to effectors (e.g. sex hormones) that diffuse through the membrane
nuclear receptor proteins
nuclear receptor proteins have (1) and (2)
- DNA-binding domain
- ligand-binding domain
The first step in response to extracellular signals is often (), which triggers downstream events that lead to transcriptional changes.
phosphorylation
An example of a multi-step cascade involves (), which is important in mammalian inflammatory and immune responses.
NF-κB
NF-κB is a heterodimer of () proteins
p50 and p65
in unstimulated cells, NF-κB is held in the cytoplasm by () -> binds to nuclear localization signal of NF-κB
I-κB
in the NF-κB signaling cascade, infection triggers activation of ()
I-κB kinase (IKK) -> I-κB is phosphorylated
during infection, phosphorylated I-κB is ubiquitinated by (1), and is thus targeted for degradation by the (2)
- E3 ubiquitin ligase
- proteasome
degradation of I-κB exposes () of NF-κB -> allows it to move into nucleus and activate transcription
nuclear localization signal (NLS)
() is where large chromosomal regions (multiple genes) are suppressed for long periods of time
transcriptional silencing
allows transcriptional silencing to persist for many cell divisions
epigenetic inheritance
transcriptional silencing is due to changes in ()
chromatin structure
chromatin structure: (1) is often transcriptionally active, while (2) is generally silent
- euchromatin
- heterochromatin
yeast chromosome III has extra copies of a and alpha genes that are not expressed:
1. a silenced copy of a is found to the right of MAT locus: (a)
2. a silenced copy of alpha is found to the left of MAT locus: (b)
- HMRa
- HMLalpha
silencing of HMRa and HMRalpha depends on () -> establish and maintain heterochromatin at HML and HMR
silencing information regulator (Sir) proteins
in silencing of HML and HMR, the ff proteins bind to these regions and recruit other Sir proteins
Abf1, Rap1, Orc1
after extra Sir proteins are recruited to HMR and HML, they spread along the chromatin and silence the genes via the ability of (1) to (2)
- Sir2
- deacetylate histones
() plays a role in establishing silencing, although its precise function is not well understood.
Sir1
Sir2 is also involved in repressing RNA pol II in the regions between the () -> helps establish heterochromatic state that prevents recombination between multiple copies of these genes
rDNA genes
() can mediate transcriptional silencing, as in IGF2 and H19
DNA modifications
IGF2 and H19 are () -> only 1 of the parental copies of a gene is expressed
imprinted genes
imprinted gene that is expressed only on paternal chromosome -> produces a growth factor needed by a developing embryo
IGF2
imprinted gene that is expressed only on maternal chromosome -> makes a non-coding RNA that may play a role in cancer.
H19
both IGF2 and H19 are found on ()
chromosome 11
regulation of IGF2 and H19 expression is due to methylation at the ()
insulator control region (ICR)
protein () can only bind to ICR when ICR is not methylated
CTCF
in the maternal chromosome, ICR is (methylated/not methylated) -> CTCF is bound to ICR and blocks enhancer to prevent transcription of ICF2 but allows transcription of H19
not methylated
in paternal chromosome, ICR is (methylated/not methylated) -> CTCF is not bound to ICR and thus H19 promoter is methylated and inhibits its transcription
methylated
Transcription is thought to be mediated by () of the DNA, and determined by access or blocking of enhancer-bound proteins.
looping
Failure of correct imprinting can lead to () – children are larger than normal at birth and are prone to cancer
Beckwith-Wiedemann syndrome
Methylated DNA can recruit specific proteins. Methyl-binding domains bind specifically to ()
methyl-cytosine
binds to methylated DNA and recruits Sin3A
MeCP2
transcriptional co-repressor that contains a histone deacetylase
Sin3A
MeCP2 represses a number of human genes – mutations in this protein can cause ()
Rett Syndrome
Rett syndrome is more common in (1) because MeCP2 is found on the (2) chromosome
- girls
- X chromosome
defective X chromosome responsible for Rett chromosome can also appear in males, but ()
since there is no extra X chromosome to mask the mutation, male fetuses don’t survive to term
