MBB 267 Week 10: Mitchel 8 Flashcards
How is transcription coupled to RNA processing?
The C-terminal domain (CTD) of RNA pol II comprises multiple copies of the serine-rich heptapeptide repeat YSPTSPS. The phosphorylation profile of the CTD varies during transcription. The different phosphorylation patterns of the CTD code act as molecular marks and allow recruitment of different processing complexes at the beginning (capping complex), middle (spliceosome) and end (cleavage/polyadenylation) of transcription in a temporally appropriate manner.
What are the ways in which splicing can recognise exon/introns?
-When are they used?
3 main ways
- Intron recognition; for small introns
- Exon recognition; for small exons
- recursive splicing; very large introns (> 50 kb)
What are the proteins involved in recognition of exon/introns?
A myriad of protein-RNA and protein-protein interactions occur, involving:
- SR proteins
- Exonic splicing enhancers (ESEs)
- U2AF
What is the difference between exon and intron recognition in splicing?
In both exon and intron recognition, U2 and U2AF are bound to each other and bind to the branchpoint. In intron recognition, the U1 complex binds at the 5’ end of intron. While in exon recognition, the U1 complex binds at the 3’ end of the 3’ exon.
How does recursive splicing work?
The intron is removed in a piecemeal manner by regenerating a functional 5’ splice site at the spliced junction.
What is alternative splicing?
-How does it work?
Most mammalian pre-mRNAs undergo multiple patterns of alternative splicing, generating distinct mRNAs from the same pre-mRNA transcript. Alternative splicing events can allow expression of distinct protein isoforms in a tissue-specific manner or within the same cell as a result of a programmed change.
-alternative splicing can regulate the expression of a single functional gene product when one pattern of splicing leads to loss of expression. The choice of which splice site is governed by the availability of alternative splicing factors, which can either activate or repress splicing at the one or other site. Splicing factors are RNA-binding proteins that recognise RNA sequences close to 5’ or 3’ splice sites, known as enhancers or silencers.
What is SMA?
How do mutations in the SMN proteins affect SMA?
spinal muscular atrophy (SMA), a debilitating muscle wasting disease that affects motor neurons that is the most prevalent genetically inherited disease in Caucasian populations. SMA is due to loss of function of the survival of motor neurons (SMN) protein
-SMN protein functions in the assembly of functional snurp particles. In humans, SMN is encoded by two genes, SMN1 and SMN2. SMA is caused by homozygous mutations within the SMN1 gene: the SMN2 sequence differs from SMN1 only by a single point mutation within exon 7. The mutation does not affect the sequence of the encoded protein but lies in an exonic splicing enhancer that is recognised by the SR protein SF2. Binding of SF2 is required for recognition of the weak 3’ splice site upstream of exon 7. Exon 7 is therefore largely omitted upon splicing of the SMN2 pre-mRNA, generating very little functional protein. The predominant splicing pattern causes exon skipping and results in a truncated and unstable protein.
How is the expression of the sex lethal gene in female specific drosophila established?
Female-specific expression of Sex lethal (Sxl) is established early in embryogenesis and then maintained through an autoregulatory positive feedback loop. This happens by excluding exon 3 in females while included in males, exon 3 is called a poison exon. Sxl also regulates other transcripts such as tra and tra2. Tra is a splice site activator that, together with the SR protein tra2, activates splicing at an alternative 3’ splice site in the doublesex transcript. In the end, both males and females produce Dsx but with different exons and therefore different functions.
What does sxl do?
Sxl directly binds to the pre-mRNA and contains two RRM domains that interact with U- rich and G/U-rich silencing elements. The RRMs also mediate protein/protein interactions with the U2 snRNP complex that form nonfunctional complexes, thereby inhibiting the ability of U2 to recognize the branchpoint.
What is trans and cis splicing?
trans = between two different RNA transcripts , and in cis = within the same transcript.
How does trans splicing work?
in trans-splicing, the spliceosome mediated splicing between one RNA containing a 5’ splice site and another transcript containing the branchpoint and 3’ splice site. The two RNAs are juxtaposed through base-pairing between sequences downstream of the 5’ splice site and upstream of the 3’ splice site
What happens to introns that lack the conserved GU and AG sequences at the 5’ and 3’ splice sites?
These are spliced by an alternative “minor” or “at-ac” spliceosome that contains variants of U1 (U11), U2 (U12), U4 (U4atac) and U6(atac). U5 is the same
