Chapter 9: Regulation of Transcription

what do recognition helices do

"read" the DNA sequences through interactions with the base pairs

what occurs in the leader sequence under high trp

- 3 and 4 interact to form attenuator and transcription stops

what are the operators for the Cro/ cI system and where are they found

- OL and OR at PL and PR promoters have three sites Cro or cI can bind to

what are coiled coils

- alpha helices wound round each other

what does RecA do

- protease that cuts cII - if cII degraded by protease, cI decreases and virus enters lytic

what is alternate pairwise cooperation

- under WT conditions, the pair binds to OR1/2 and OR3 only binds at much higher cI conditions - when OR1 mutated, pair binds OR2/3 - binding at OR1/2 turns off PR and on PRM

how can regulation of gene expression be fine tuned

regulator abundance, localization, phosphorylation

how many transcriptional regulators control yeast cell type and what are the cell types/ their traits

- 4 transcriptional regulators - a and alpha cells are haploid, can mate with opposite cell type - a/alpha cells are diploid and result from the mating of alpha and a cells, cannot mate but undergo meiosis when starved

what happens if cII is not produced in high enough quantities

- Cro binds to PRM promoter, represses cI production ensuring expression of lytic genes - levels of cII are thus crucial to whether the cell enters lysogenic or lytic phase

how is galactose metabolism regulated

- Gal4 regulates genes that are responsible for galactose metabolism - gal4 activates transcription by binding to uasg sequence - gal4 activity is regulated by gal80 and gal3 which respond to galactose in the cell - in the absence of galactose gal80 binds to gal4 preventing transcription - in the presence of galactose, gal3 binds to gal80 preventing gal80 from binding to gal4 and allowing gal4 to recruit SAGA and mediator to activate transcription

how can DNA modifications help silence genes

- IGF2 makes a growth factor needed in embryo dvlpmt, H`9 makes noncoding RNA that may play role in cancer. both are found on c-some 11 - IGF2 on maternal, H19 paternal due to methylation on insulator control region. the protein CTCF can only bind to ICR when ICR is unmethylated as is in the maternal c-some - binding of CTCF means transcription of H19 and blockage of IGF2 - w unmethylated paternal c-some, IGF2 can be transcribed and H19 cannot due to methylation

what is MeCP2

- binds methylated DNA and recruits Sin3A, a transcriptional co-repressor that contains histone deacetylase - mutations here can cause Rett Syndrome

how does lysogenic phase get promoted by cI and Cro

- cI binds to OR1 and OL1 first, recruiting more cI to bind to the weaker OL2 and OR2 sites - cI oligomerizes to form a looped out structure. this stops RNA pol binding to PL and PR and promotes transcription of PRM leading to increase cI levels - more cI means binding to OR3 and OL3 stopping activation of PRM and suppresses cI synthesis (cI autoregulates) - if there is insufficient cI, Cro accumulates and binds with greatest affinity to OR3 blocking PRM and suppressing cI synthesis. more Cro synthesis leads to binding at OL and OR sites autoregulating Cro

what are allosteric effectors and what is an example

- estrogen - binds to regulatory proteins and changes their conformation

how does ume activate or repress transcription

- ume6 binds DNA when there is enough N and C in the cell recruiting co-repressors Sin3, Rpd3, and Isw2 - Rpd3 is a histone deacetylase which promotes more compact chromatin and represses transcription - isw2 is a nucleosome remodeling enzyme which helps establish altered chromatin pattern - ume6 Is phosphorylated in the absence of N and c causing dissociation of these supplemental proteins and addition of a coactivator called Ime1

how does chromatin structure influence eukaryotic transcription

hyperacetylated chromatin tends to be actively transcribed, whereas levels of transcription are low in hypoacetylated chromatin

what are eukaryotic genes most often controlled by

- distal regulatory sequences, called enhancers - enhancers can be upstream or downstream

what do two component systems contain and how do they work

- sensor kinase and response regulator - histidine in the sensor becomes autophosphorylated on receipt of signal - phosphoryl group is then passed onto aspartic acid residue in a response regulator protein

how are some RNA pol holoenzymes activated

- sigma54 requires ATP hydrolysis to drive promoter opening

what is combinational control

response of a gene to multiple signals and regulatory proteins

what four proteins regulate yeast cell type and what locus are they found on

- MAT locus on chromosome III - a1, alpha1, alpha2, MCM1 - MCM1 activates transcription of a-specific genes - alpha1 activates transcription of alpha specific genes - alpha2 represses a-specific gene transcription

what proteins interact with beta sheets and loops

- MetJ repressor - p50 and p65 which are part of the mammalian transcriptional regulator nuclear factor NF-KB are largely made of beta sheet - immunoglobulins are entirely made of beta sheets and connecting loops

what is the NF-KB pathway

- NF-KB is made of p50 and p65 - in unstimulated cells, NF-KB is kept in the cytoplasm by I-KB - I-KB binds to the region on NF-KB that targets it to the nculeus - infection triggers a cascade that activates the I-kB kinase and I-KB is phosphor - phosphor I-KB is ubiquitinated by e3 ubiquitin ligase and thus targeted for degradation by the proteasome - this exposes the NLS on NF-KB and allows import. into nucleus to activate transcription

what is Rett syndrome

- a progressive neurological disorder that only occurs in females. - similar to autism - mecp2 on x chromosome, so males only need one copy to be lethal, females have one good and one mutant

what occurs in alpha/a diploid cells

- a1 and alpha2 form heterodimer - this dimer binds different DNA sequences to the monomers and represses haploid specific genes - alpha1 and RME1 (represses meiosis genes) are suppressed - lets meiosis occur

what is NtrC

- activator that activates genes that are important for nitrogen metabolism - binds at enhancer elements and activates transcription when phosphorylated - phosphorylation triggers oligomerization of NtrC and promotes interaction with the polymerase at the promoter - oligomerization also stimulates its rate of ATP hydrolysis which helps promote the formation of the open complex

what is CAP and how does it regulate

- also called cAMP - catabolite activator proteins - glucose depletion leads to increase in cAMP levels which binds to CAP and increases binding to DNA - CAP enhances RNA pol by binding to CTD

what are bZIP and bHLH proteins

- bZIP: basic region-leucine zipper proteins have long alpha helices of about 60 amino acids. the two helices have hydrophobic leucine residues on the surfaces that zip together. at the N terminal end, the helices splay and sit in the DNA major groove. when not bound to DNA, the basic region is disordered. - bHLH: basic region helix loop helix proteins also become structured upon binding to DNA

what is the difference between class I and II promoter for CAP

- class 1: CAP binds upstream of promoter - class 2: CAP binding site overlaps with the RNA pol binding site

what are lysis and lysogeny regulated by and how does this occur

- cro, cI, cII - these bind to sites in the phage chromosome that control transcription at the four promoters: PR, PL, PRE, PRM - PR promoter first produces Cro and cII - if there is enough cII, it stimulates transcription of PRE which produces cI - cI stimulates PRM which makes more cI. High cI levels bind to PR and PL and repress lytic genes and activates transcription of integrase therefore promoting lysogeny

how are HMLa and HMRa silenced

- depends on silencing information regulator (sir) - establish and maintain heterochromatin at HML and HMR - other proteins bind these regions (Abf1, Rap1, Orc1) which recruit other Sir proteins - extra sir proteins spread along chromatin and silence via the ability of sir2 to deacetylate histones - genes close to the yeast telomere are silenced in a similar way - sir2 is also involved in repressing rna pol 2 transcripts in the regions btwn the rDNA genes- this helps establish chromatin in which recombination is prevented btwn these genes

what is the leader sequence in trp operon and what does it contain

- has intrinsic terminator/ attenuator - leader sequence has 4 blocks: 1,2,3,4 that can form alternative pairing arrangements. - 1 and 2 can form a stem loop - 2 and 3 can form a rho-independent terminator - 2 and 3 can also form a stem loop

what is the trp repressor

- helix turn helix protein - trp repressor binding to the DNA depends on the level of tryptophan in the cell - trp repressor can only bind when it is bound to tryptophan which only occurs under high tryptophan levels

what is a common eukaryotic DNA binding domain

- homeodomain - monomeric helix-turn-helix - helix 3 fits in the major groove and side chains interact with DNA base-pairs - the N terminal arm makes contacts in the minor groove

what are LCRs and where are they found

- locus control regions - found in more complex eukaryotes - LCRs have a combination of enhancer and insulator elements - beta-globin genes are controlled by a locus control region

how does ELK1 work with mitogens

- mitogens are extracellular signals that promote entry into mitosis - ELK1 recruits mediator - in the absence of mitogens, ELK1 binds to serum response factor but doesn't activate transcription - mitogen binding at the cell surface activates kinases which phosphorylate ELK1. phosphorylated ELK1 recruits mediator promoting transcription

regulatory proteins are often _________ and have ___________ that recognize specific sequences

- modular: have extra domains that aid oligomerization, activate or repress transcription, interact with other regulators - DNA binding domain

how is human interferon beta regulated

- multiple proteins bind the interferon to form an enhanceosome - bindings aided by an architectural DNA binding protein called HMG-I(Y) - HMG-I(Y) includes DNA bending which helps other factors bind. the bend is removed in the final complex

how does the trp operon work

- no trp: trp repressor cannot interact with DNA and prevent RNA pol from binding meaning transcription of trp genes - w trp: allosterically changes conformation of trp repressor, binds promoter and shuts off/ prevents RNA pol binding

what occurs in the lytic phase

- production of lots of copies of the virus which eventually leads to cell lysis and allows the phage to escape a cell that may not survive

what is a zinc finger

- prominent part of many DNA binding domains - has domain of ~30 amino acids with an alpha helix and two beta strands wrapped around a central zinc ion - zinc interacts with two cysteines and two histidines - proteins have several zinc fingers each of which inserts its alpha helix into the minor groove - zinc fingers are the most common DNA binding domains in the human genome

what does DNA damage do

- prompts the host SOS response, activating RecA - RecA cleaves cI so it cannot dimerize or cooperate with other cI units - cI dissociates from operator sites allowing transcription from PL and PR and lytic growth

what is the B subtilis adenine riboswitch

- regulates adenine synthesis and transport - gene expression depends on whether a terminator or anti-terminator forms - in low adenine, the RNA structure has regions 2 and 3 forming an anti-terminator - in high adenine, regions 3 and.4 form a terminator

what occurs in the leader sequence under low trp

- ribosome stalls when translating leader peptide - region 1 is blocked so 2 and 3 form the stem loop preventing 2 and 3 from binding and forming a terminator allowing transcription to continue

how does heat shock response work

- rna pol sometimes pauses after transcribing the first few dozen nucleotides. this is used to prime hsp70 for rapid transcription in response to heat shock - without heat shock, GAGA factor binds upstream of hsp70 and recruits NURF which keeps the promoter free of nucleosomes and allows polymerase binding for quick synthesis of hsp70. transcription starts but at 25 degrees celsius pauses because the complex is not phosphorylated enough - when heat is present, hsf trimerizes and binds to heat shock elements. hsf then recruits mediator to the promoter as well as pausing complexes anda. kinase which phosphorylates the CTD and allows transcription to resume

how does HIV use anti-termination

- tar site forms a stem-loop called the tar element which leads to termination unless tat protein present - tat viral protein binds to tar and phosphorylates rna pol CTD and several other proteins - phosphor of CTD relieves pausing so prevents premature transcript formation

what is anti-termination in phage lambda

- transcription from PL and PR promoters successfully makes Cro and N but encounters termination sites tL and tR terminating the transcripts - when N protein builds up, it stops termination - N protein binds to nut sites in the transcribes RNA in a step-loop form. Several proteins are recruited and interact with RNA pol allowing it to transcribe through tL and tR - with termination at tL and tR suppressed, downstream genes can be transcribed including Q - Q protein overrides termination at the PR' promoter allowing transcription of late lytic genes - Q prevents termination by binding DNA. the Q binding site (qbe) is btwn the -10 and -35 of the PR' promoter - without Q, transcription terminates very early just downstream of -10 - Q directly contacts sigma subunit (sigma 70 for this gene) and allows elongation to resume - Q remains associated with elongation complex

what occurs in lysogeny

- virus integrates into host genome and becomes dormant - integrated virus is called a prophage - virus multiplies in conditions where there may not be enough cells nearby to infect

how do CAP and the LacI repressor regulate the lac operon

- when low glucose and low lactose, CAP binds and enhances transcription of lac genes. lac repressor doesn't bind. - when high glucose and low lactose, lac repressor doesn't bind but neither does CAP so weak expression of lac genes. - with no lactose, regardless if glucose is high: lac repressor binds and there is no transcription

binding affinities of what two proteins at ______ DNA binding sites determines lysogenic or lytic stage

Cro and cI, six

what happens if the operator is distal to a gene

DNA must loop around for the regulatory protein to interact with the polymerase, sometimes aided by architectural DNA binding proteins

what are the two regions of a riboswitch

aptamer that binds to the metabolite and an expression platform which controls transcription or translation

what did Dodd find

cI binding to OR3 is helped by an interaction with OL3 likely due to DNA looping so cI can interact with both operators at high cI conditions

what is a nuclease protection assay in the context of cI binding

cI incubated with OR1, OR2, OR3 - cI interacts w the gel then nuclease added - little protein needed to see binding with OR1, more w OR2, even more still w OR3 - when only N terminal domain of cI is present, no cooperative binding to OR1 and OR2, affinity for OR2 lower - loss of OR1 had no effect on binding affinities for O2/3 when only N terminal was present

how many domains do Cro and cI have

cI: two, can form tetramers and oligomers Cro: single small domain that binds DNA as a dimer

what does MerR do

compresses helix to change the way promoter elements are spatially related to each other - can change large space of 19bp to 17bp (normal) which enhances transcription

what is beckwith-wiedemann syndrome

failure of correct imprinting can lead to larger children who are prone to cancer

what are operators

found in bacteria; sequences recognized by regulators, typically found close to the promoter or overlapping it

many genes are regulated by proteins that bind ______ the gene, most often _______ of the promoter

near; upstream

an co-activators and co-repressors bind DNA alone

no; they are recruited by specific regulatory proteins

what are riboswitches

portions of a transcript that can directly bind a small molecule that controls the RNA secondary structure, regulating transcription or translation

what is anti-termination

some phage lambda genes are transcribed only when transcription termination is actively prevented

DNA binding motifs are

tailored to fit DNA (helix-turn-helix forms dimers with recognition helices spaced 3.4nm apart to fit into neighboring grooves)

what are TAL proteins and what was found about them

tal effectors activate host genes that help bacteria thrive - have tandem repeat of 34 amino acid sequence - bonas noticed that AA sequence highly conserved - stoddard looked at crystal structure of TAL proteins bound to DNA - TAL repeats arranged in right handed spiral that follows major groove of DNA - each tal repeat forms a pair of alpha helices - tal being used for engineering of proteins w specific binding affinities

what is transcriptional silencing

where large chromosomal regions are not transcribed for long periods of time, perhaps many cell divisions