Eukaryotic gene expression Flashcards
How many RNA pols in eukaryotic?
3.
I
II
III
What does RNA pol I transcribe for?
rRNA
What does RNA pol II transcribe for?
mRNA (via hnRNA)
What does RNA pol III transcribe for?
tRNA (and other small RNAs)
What are the general factors that interact with RNA pol II?
Form basal transcriptional apparatus along with RNA pol II.
What are the upstream factors that interact with RNA pol II?
DNA-binding factors recognising short consensus sequences, not regulated and required for adequate transcription.
What are the inducible factors that interact with RNA pol II?
as upstram factors, but regulated to provide control of gene expression in eg stimulus specific manner.
What are the general factors known?
(TFII)D, A, B, F, E, H
combine with RNAPII to start basal transcription apparatus.
What is used to recognise promoters?
TFIID = TBP + TAFs
= TATA-binding protein + TBP-associated factors
Positions RNAPII in right place.
Only general initiation factor which binds promoters in sequence specific manner.
Binds to DNA in minor groove.
Bends DNA by 80 degrees.
What happens to CTD of RNA pol II?
Reversibly phosphorylated, allowing promoter release and coordinating post-transcriptional events.
What are cis-acting control elements (DNA)?
Promoters
Enhancers/repressors.
What are trans-acting transcription factors (proteins)?
Consitutively expressed or induced.
2 domains - DNA binding and activation domains.
Where do down mutations happen?
3 locations corresponding to 3 elements which make up promoter.
GC box -90
CAAT box -75
TATA box -30
What is needed for transcription to be activated?
Particular combo of control elements can activate transcription only when the appropriate activator proteins are present (and potentially expressed).
What do DNA binding domains do?
Recognise target sequences in enhancers or promoters.
What do activation domains do?
Function to influence transcription by contacting basal apparatus.
Where does the transcriptional co-activator bind?
Does not bind DNA itself, but bridges btwn activator and basal transcription apparatus.
What controls gene expression?
Cis-acting (consesus) sequences and trans-acting (transcription) factors in combinational fshion.
How do transcription factors uses 2 domains?
bind upstream of tsp, contacting basal transcriptional apparatus (via mediator) to increase transcriptional initiation.
What is chromatin?
the material of which the chromosomes of organisms other than bacteria (i.e. eukaryotes) are composed, consisting of protein, RNA, and DNA.
What is a nucleosome?
a structural unit of a eukaryotic chromosome, consisting of a length of DNA coiled around a core of histones.
What are the two types of nucleosome remodellers?
2 main classes:
ATP-dependent nucleosome remodellers.
Modifiers of histone acetylation
What does acetylation/deacetylation?
HAT/HDAC; acetylation both alters nuclesomal structure and recruits bromodomain containing proteins
What happens in pre-mRNA processing?
- Transcription, 5’ capping (at 25-30 nt of elongation)
- Cleavage at Poly(A) site
- Polyadenylation
- RNA splicing
What bits are 100% conserved in exon-intron junctions?
GU
Branch point (A)
What is lariat formation?
Lariat formation involves cleavage of the precursor at the 5’ splice site (5’ SS) and the formation of a 2’-5’ phosphodiester bond between the guanosine residue at the 5’ end of the intron and an adenosine within the intron. Intron AKA lariat separates and allows exons to join together.
How many U-rich small nuclear RNAs (snRNAs) are involved in splicing.
5 U1,2,4,5,6
WHat does the U2 RNA do?
Pairs in such away that the A of the branch sticks out, allowing it to take part in the first transesterification reaction.
How does splicing of pre-MRNA occur?
snRNPs assemble on pre-mRNA to form a spliceosome.
1st U1, U2 bind.
then 4, 5 and 6 enter as a complex.
Spliceosome rearranges - release 1, 4 - transesterification occurs.
2nd rearrangement leads to 2nd transesterification to occur.
Excised intron and snRNPs dissociate.
What do -ve factors do to U1 and U2?
Prevent binding or block spliceosome assembly after U1 or U2 binding
What do +ve factors do?
Enhance spliceosome assembly at sites that are poorly recognised
What needs to happen to CTD for initiation to occur?
Phosphorylation
Do mRNA have diff half lives?
Yes diff stabilities.
short = regulatory proteins - AU rich in 3’-UTR (AUUUA)
long = structural proteins/ key metabolic enzymes
What does 5’ cap do?
Aids mRNA nuclear export
Protects from 5’-3’ exonuclease degradation in cytoplasm.
Enhances translatability of transcript
What does poly-A tail do?
Participates in termination of transcription.
Aids mRNA nuclear export
Protects from 5’-3’ exonuclease degradation in cytoplasm.
Enhances translatability of transcript
Is mRNA exported?
Yes it is exported by mRNA exporter.
Leaves via pore of nucleus
What can regulate mRNAs in cytoplasm?
Proteins that bind to it - regulates translation and degredation.
