L3, Eukaryotic Transcription Flashcards
+ Name the archaeal superphylum propose to be an archaeal ancestor to eukaryotic lines, give one example, which theory does this relate to in phylogenetics?
- ‘Asgard Superphylum’
e.g. Lokiarchaeota, Heimdall (played by Idris Elba) - Two-domain tree of life
+ Why is lokiarchaeota of particular interest to phylogeneticists?
It has been shown to be more closely related to eukaryotes than any other prokaryotic lineage
Types of RNA P in eukaryotes:
- Type I: Found in nucleolus, produced rRNA
- Type II (KEY): Found in nucleolus; produces mRNA and snRNA
- Type III: Found in nucleolus, produces rRNA, tRNA and snRNA
- Separation of function between types
Overview of transcription initiation in eukaryotes:
- Does not involve sigma factor by a collection of proteins instead (basal/general transcription factors)
- Main interaction complex is mediator (CRSP/MED), coordinates interaction between activators and basal TFs
- Also important to consider epigenetics; chromatin structure must be relaxed (euchromatic) in order for transcription, with temporary displacement of histones)
How large is the human genome and how many genes does it contain?
- ~3 x 10^9 bp
- ~30k genes
Post-transcriptional processing in eukaroytes:
- Addition of 5’ cap
- Addition of poly(A) tail
- RNA splicing (introns removed)
- (Eukaryotic genes are monocistronic as a rule)
Basic structure of RNA P in euk vs prok?
- Basic crab claw shape of the core is conserved between archaea (crucially, asgard archaea), bacteria and eukaryotes
Describe the features of archaeal RNAP: (compare to eukaroytic)
- Characteristic protruding stalk (made up of a heterodimer of E and F subunits)
- This heterodimer is found across all 3 classes of eukaryotic RNAPs
- The stalk functions to interact with nascent RNA transcript
- Stimulates transcription processivity
- Recruits RNA-processing factors in eukaryotes
Structure of RNA PII promoters: (conserved regions)
- Inr region: found at startpoint; Py2CAPy5
- TATA box: A-T rich octomer 25bp upstream of start-point -> recognised by TBP
- BRE element: interacts with general transcription factor TFIIB
- DPE: Downstream promoter element (found in TATA-less promoters)
- -> These sequence elements all function by binding regulatory proteins
In what order do TFs bind to form transcription initiation complex?
- TFIIA and TBP subunit of TFIID bind
- TFIIB binds
- TFIIF + RNA PII bind
- TFIIE binds and forms a complex with TFIIF (Pre-IC formed)
- TFIIH binds (IC formed)
What is the relation between TFIID and TBP? What is the role of TBP?
- TBP is actually a subunit of TFIID, the largest general transcription factor
- It is stabilised by TFIIA
- The TBP subunit has a characteristic saddle-shape, inducing a ~90 degree bend at the TATA box
What is the role of TFIIH?
- Unwinds the initiation site
- Also phosphorylates of the Pol II-CTD
Why is a pre-initiation complex useful?
- It is an inactive, preassembled precursor to the transcription initiation complex
- Allows for streamlined reaction to external stimuli -> transcription is quick
As well as TBP, what other features does TFIID have? What is their role?
- At its core, TFIIF has 8 to 16 TBP associated factors (TAFs)
- Whilst TBP is sufficient for basal transcription on its own, these TAFs also influence interaction with downstream regulatory factors
Features of TFIB: Size, binding site, key domains
- 35 kDa
- Binds to BRE
- Has a zinc-binding N-domain which interacts with Pol II and TFIIF allowing recruitment of Pol II
- C domain interacts with DNA, TBP and Pol II
- The linker between these two displays limited homology to part of bacterial sigma factors
