Molecular Biology - RNA Polymerases PDF

BIOL10221 “Molecular Biology” Dr David Boam Module 4 - From DNA to RNA Lecture 2: RNA polymerases  Aims of this lecture Aims To show that there are differences in complexity and organization between the prokaryotic and eukaryotic transcriptional apparatus RNA po...

BIOL10221 “Molecular Biology” Dr David Boam Module 4 - From DNA to RNA Lecture 2: RNA polymerases  Aims of this lecture Aims To show that there are differences in complexity and organization between the prokaryotic and eukaryotic transcriptional apparatus RNA polymerases in prokaryotes and eukaryotes RNA polymerase-associated general factors Lecture 4/2 RNA polymerases  Transcription initiation by prokaryotic RNA polymerase (Sense) b  a Coding Strand +1 s 3’ 5’ -35 -10 3’ 3’ 5’ Template Strand (Antisense) 5’ RNA Polymerase holoenzyme sigma (s) – promoter recognition 2 x alpha (a) – assembly, activation 2 x beta (b+b’) – catalysis, termination Omega () – assembly, folding, required for some genes Lecture 4/2 RNA polymerases  Transcription initiation by prokaryotic RNA polymerase s (Sense) Coding Strand 5’ b 3’ -35 -10  3’ 5’ Template Strand (Antisense) s is released and the core enzyme (aabb’) is competent for elongation Lecture 4/2 RNA polymerases  RNA Polymerases Prokaryotic One RNA polymerase Eukaryotic Genes which make different types of RNA have Three RNA polymerases – Different architecture Pol I - ribosomal RNA genes (rRNA) – Use different RNA polymerases Pol II - protein coding genes – Each type of RNA (mRNA) polymerase needs a small nuclear RNA (snRNA) distinct set of accessory factors Pol III - transfer RNA (tRNA) rRNA and snRNA Lecture 4/2 RNA polymerases  Assembly of the RNA polymerase II initiation complex -part 1 TFIID IIA TBP IIB  TFIID = TATA binding protein (TBP) + TBP-Associated Factors (TAFs)  Only specific DNA-protein interaction  TBP has sequence similarity to s  TFIIA - Helps TFIID to bind.  TFIIB - sets distance from TATA element to start site Lecture 4/2 RNA polymerases TBP-DNA complex From underside note b sheet surface contact with DNA minor groove Head on view down DNA barrel Saddle structure Lecture 4/2 RNA polymerases TBP-TFIIB-DNA complex Lecture 4/2 RNA polymerases TBP-TFIIA-DNA complex Lecture 4/2 RNA polymerases  Assembly of the RNA polymerase II initiation complex -part II RNA polymerase II 10 subunits IIH IID IIJ IIA IIE IID IIF IIB Pol II TFIIE,F,H,J,K  Interaction with D-A-B complex and recruitment of polymerase II  Blocks non-specific binding of polymerase II to DNA (like σ)  Promoter clearance  Helicase  Processivity, elongation  Transcription-coupled DNA repair Lecture 4/2 RNA polymerases  RNA Polymerase I UCE core SL1 = TBP+ pol I-specific TAFs UBF RNA TBP UBF Pol I Upstream binding factor (UBF) binds to UCE and core element in DNA RNA Polymerase I binds to complex and transcription is initiated Lecture 4/2 RNA polymerases  RNA polymerase III – tRNA genes TFIIIC TBP RNA pol III A box B box TFIIIB Promoter within coding region TFIIIC binds to B box TFIIIB recruited upstream of start site – TFIIIB = TBP + 2 pol III-specific TAFs Once TFIIIB is recruited, TFIIIC is dispensable RNA polymerase III recruited and initiates transcription Lecture 4/2 RNA polymerases Other RNA polymerase III genes TFIIIC TFIIIA 5S rRNA genes are similar to tRNA genes but use an A box C box extra factor TFIIIA to recruit TFIIIC snRNA genes have a similar structure to pol II genes PSE TATA Upstream promoter Proximal TATA box (probably Sequence Element recognised by TBP) Lecture 4/2 RNA polymerases  Summary There are differences in complexity and organisation between the prokaryotic and eukaryotic transcriptional apparatus RNA Polymerases are macromolecular complexes Prokaryotes have 1 polymerase, eukaryotes have 3 Eukaryotic polymerases – specialised, gene type-specific – use general transcription factors Lecture 4/2 RNA polymerases  Intended learning objectives You should know that Keywords RNA Polymerases are macromolecular RNA polymerase complexes there are differences in complexity and Recruitment organisation between the prokaryotic and Multi-protein complex eukaryotic transcriptional apparatus eukaryotic RNA polymerases are gene - Sigma type specific TBP Understand the process of transcriptional TFIIA – J initiation in prokaryotes and appreciate that the process is similar in eukaryotes tRNA promoter Be able to identify and understand the rRNA promoter function of general transcription factors in the function of eukaryotic RNA mRNA promoter polymerases Lecture 4/2 RNA polymerases