Post-Transcriptional Regulation Flashcards
What is RNAPII involved in?
capping, splicing and polyadenylation coupled to transcription
What does phosphorylation of the CTD of RNAPII do?
release RNAPII from the promoter to start RNA elongation which allows different sets of proteins to bind for capping, splicing and polyadenylation
What is mRNA capping?
5’ to 5’ addition of GMP to the 5’ end of the RNA transcript
When is the capping reaction started?
when an RNA is synthesised to ~25nt
What are the roles of capping?
- signals translation initiation by ribosome binding
- ensures correct processing and export of mRNA through binding a cap binding complex
- stabilises and protects 5’ end of mRNA from degradation
What are the 3 steps of capping?
- phosphatase removes a phosphate from 5’ end of primary transcript
- guanylyl transferase adds a GMP in a reverse linkage (5’-5’ instead of 3’-5’)
- methyl transferase adds a methyl group to the guanosine
What are all 3 enzymes for RNA capping associated with?
the CTD
What are the 3 splicing consensus sequences?
- 5’ GU - donor site
- 3’ CAG - acceptor site
- branch point - loose consensus
What are the 2 basic steps of RNA splicing?
- join the 5’ end of intron to a branch point A to form a lariat loop
- cut the 3’ end of intron and join the 2 exons
What type of reaction happens in RNA splicing?
transesterification
What are the 5 snRNAs and what are their functions?
- U1 - recognises 5’ splice site
- U2 - initial binding to branch site
- U4 - sequesters U6
- U5 - brings 2 exons together
- U6 - catalyses 2 transesterification reactions with U2
How are snRNPs formed?
each snRNA is complexed with more than 7 protein subunits
What does the spliceosome do?
- recognise the intron splice sites
- bring the two ends of the exon together
- remove the intron
What are the 4 steps of RNA splicing?
- U1 forms base pairs with the 5’ specific junction and the BBP and U2AF recognises the branch-point site
- U2 displaces BBP and U2AF and forms base pairs with the branch-point site consensus sequence
- U4/U5-U6 triple snRNP enters the reaction and once the U4/U6 base pairs are separated, U6 displaces U1 at the 5’ splice junction which creates the active site that catalyses the first phosphoryl-transferase reaction
- additional RNA-RNA rearrangements create the active site for the second phosphoryl-transferase reaction, which then completes the splice site
What does U1 RNA contain?
- several distinct stem-loop domains
- 8 core Sm proteins and 3 U1-specific proteins
What are Sm proteins?
8 common proteins constituting the core structure of snRNPs
What is Sm binding site required for?
interaction with common snRNP proteins
What is recognition of consensus sequences important for?
to ensure the accuracy of the position for RNA splicing
What are the 2 common types of splicing errors due to imperfect RNA pairing?
- exon skipping
- cryptic splice-site selection
How can splicing errors be minimised?
- coupling splicing to transcription to avoid exon skipping
- exon definition
What is the exon definition hypothesis?
the exon is bound by a group of SR proteins that serve as splice enhancers; they recruit U1 and U2 to define 3’ and 5’ exons respectively
What are SR proteins?
proteins rich in Ser and Arg
How are introns packaged into complexes?
by hnRNPs (heterogenous nuclear ribonucleoproteins)
What does abnormal processing of the β-globin primary RNA transcript in humans produce?
aberrant β-globins causing β-thalassemia disease
