BIOL 300 - Chapter 18

what is the difference between prokaryotic and eukaryotic transcription in template?

prokaryotes have no chromatin structure, but eukaryotes have chromatin structure and must be in an open state for transcription, moving/repositioning nucleosomes if necessary

what is the difference between prokaryotic and eukaryotic transcription in promoter recognition?

prokaryotic promoter is conferred by sigma factor as a part of the polymerase holoenzyme, while eukaryotic promoters are recognized by basal/site specific transcription factors

what are transcription factors?

accessory proteins that promote/repress RNA polymerase from binding to DNA - don’t incorporate into poly holoenzyme, and dont directly bind to transcription site

what is the difference between prokaryotic and eukaryotic transcription in polymerases?

prokaryotes only have one RNP, eukaryotes have three polymerases; RNA Pol 1, RNA Pol 2, RNA Pol 3

what is the purpose of RNA Pol 1?

transcribes bigger rRNAs

what is the purpose of RNA Pol 2?

transcribes mRNA and some small RNAs like microRNA

what is the purpose of RNA Pol 3?

transcribes tRNA, 5S rRNA, and other small RNAs like snRNA

what’s the difference between prokaryotic DNA Poly III and eukaryotic RNA Poly III?

prokaryotic DNA Poly III isn’t a polymerase, it’s a replicase, but eukaryotic RNA Poly III will produce mRNA.

what is the difference between prokaryotic and eukaryotic transcription in termination?

prokaryotes have terminator sequences, eukaryotes have a cleavage sequence instead

what is a CTD?

C Terminal Domain

what is the purpose of promoters?

position polymerases to the transcription start site

what types of promoters do RNA Pol I and Pol III use?

constitutively active promoters - limited number of elements and TFs that bind to them, and rRNA and tRNA are needed for all stages of cell cycle

what types of promoter does RNA Pol II use?

some genes need constitutive expression, others need expression in specific conditions; requires many control elements and the TFs that bind to them are needed for recognition of sequence elements

what are the two elements of Polymerase I Promoters?

core promoter, and upstream promoter element

what are the features of the Pol 1 core promoter?

sufficient for transcription at moderate levels, GC rich and only AT rich at start point, bound by SL1 complex containing TBP

what are the features of the Pol 1 upstream promoter element?

not absolutely necessary for transcription, just significantly increases the rate; GC rich and bound by upstream binding protein

what is the role of the upstream binding protein?

stimulates SL1 binding that wraps DNA to bring UBP close to core promoter, prevents H1 binding and inactive chromatin

what are the three types of Pol III promoter?

type 1 (5s rRNA), type 2 (tRNA), type 3 (snRNA)

what are the properties of Type 1 Pol III promoter?

has box A and box C, and a fixed downstream/internal promoter. TFS that bind to downstream promoters recruit TBP to upstream posititon to recruit polymerase III

what are the properties of Type 2 Pol III promoter?

internal/dowmstream, has box A and B, has variable positioning in respect to the transcriptional start site. TFS that bind to downstream promoters recruit TBP to upstream posititon to recruit polymerase III

what are the properties of Type 3 Pol III promoter?

upstream like polymerase II, has Oct, PSE (both allow binding of additional factors to enhance transcription), and TATA box (necessary and sufficient)

what are the two functions of the TBP (TATA box binding protein)?

binds to minor groove of TATA box and binds/recruits RNAP to promoter

what are the two ways to position TBP?

in the TATA containing promoter (involves half of pol 2 and pol 3 type 3) - binds directly to the TATA box, and in TATA-less promoters (recruited by TAFs (TBP associated factors) that bind to promoters)

what are the features of Pol 2 regulatory sequences?

used for mRNA and microRNA, contains a promoter made up of the core promoter and upstream promoter elements (core promoter isn’t enough)

what makes up the polymerase II core promoter?

initiator sequence, a TATA box (-25 bp)/DPE (downstream promoter element) (+32 bp), binds to general/basal TFs

what are the features of the polymerase II UPE?

closer to start point, major contribution to promoter specificities, and is responsive to the GC box, CAAT box, Oct, etc

what are the three regulatory cis elements?

enhancer, silencer, insulator

what is the purpose of the enhancer?

similar to upstream promoter - increases expression level (distance and orientation doesn’t matter)

what is the purpose of the silencer?

transcription repressor that decreases expression (distance and orientation doesn’t matter) - less abundant than enhancers because the default state of transcription is already inactive

what is the purpose of the insulator?

prevents enhancers from active, and stops heterochromatin from spreading

what are the two types of transcription factors?

basal and general

what is the role of basal transcription factors?

bind to core promoter to position polymerase - needed but not enough alone, because they don’t provide specificity

what is the role of transcription factors?

work as activators - required for normal expression in vivo, and bind to promoter/enhancer to respond to specific conditions

what are the three stages of RNA poly 2 initiation?

pre-initiation (closed) complex, open complex, and promoter escape

what occurs during the pre-initiation (closed) complex phase in RNA poly 2?

BTFs and pol 2 pind to the core promoter, and the BTS recruit everything that needs to come to the promoter in their specific order (TF2D (made up of TBP and TAFs) to bind to the TATA box, then TF2B, F, E, and H (TF2A can come whenever))

what occurs during the open complex phase in RNA poly 2?

2 basal TFs join (TF2J and TF2H) to melt the DNA open

what occurs during the promoter clearance/escape

serves as the transition to elongation, and involves the release of most TFs. Requires phosphorylation of pol 2 CTD tail TF2H, and is the only chance for abortive initiation

what are the features of the Pol 2 CTD (C-terminal domain)?

has multiple repeating peptide units with a consensus sequences of 7 amino acids: (YSPTSPS), and is regulated by phosphorylation of serines and threonines

what tare the eukaryotic transcription elongation events?

navigate through the nucleosome (its disassembled until polymerase passes then reassembles after), and is paired with RNA processing AS transcription occurs

what are the steps of transcription?

RNA poly 1 enters w/o phosphorylation and is then phosphorylated, poly 2 is initiated and elongation starts (with splicing simultaneously occuring), dephosphorylation, then binding of phosphorylation factors to the phosphorylated areas that still remain, then more dephosphorylation

how does transcription termination occur?

transcription occurs downstream for over 1.5kb and eventually falls off since there’s no terminator, but there’s a signal for cleavage of pre-mRNA for the polyA tail through polyadenylation by enzymes bound to the CTD of RNA pol2, so anything after that point is spliced off