RNA splicing Flashcards
Is splicing co or post-transcriptional?
Cotranscriptional
What is the function of splicing?
To remove introns and join exons together
What are the 3 key highly conserved sequences needed for splicing?
- A 5’ GU splice site
- A branch site (A)
- A 3’ AG splice site
What carries out splicing?
The spliceosome
What is the spliceosome?
Complex of proteins and snRNA that assemble into ribonucleoprotein complexes (snRNP), then the snRNPs assemble into the spliceosome
What is snRNA? What does it do?
Short nuclear RNA. Are catalytic ribozymes that complementary base pair to the pre-mRNA and do the cutting and pasting
What are snRNP proteins? What do they do?
Proteins that assist the snRNA, but don’t do any splicing. They position the snRNA so it can catalyze splicing and will remodel the entire spliceosome during splicing
What is alternative splicing?
Where a single gene can be spliced in different ways to produce multiple transcripts and multiple polypeptides from the same gene
Is alternative splicing common?
Really common. 95% of mammalian genes
How does alternative splicing occur?
Mediated by RNA binding proteins that enhance or suppress the selection of a particular splice site by the spliceosome
What is the advantage of alternative splicing?
Allows multiple proteins to be encoded by the same gene, so the organism needs fewer genes
What are the 3 genes involved in the sex determination pathway in Drosophila?
Sxl, tra, and dsx
How does alternative splicing leads to the development of female Drosophila?
- A TF early in the development of females activates expression of sxl
- sxl gets activated again later, and the sxl that was already around changes splicing so that exon 3 gets spliced out and the protein is fully functional
- SXL alters the splicing of tra mRNA to produce a fully functional protein
- TRA will alter the splicing of dsx so that it contains exons 1,2,3 and 4. This version of the TF binds to male sex differentiation genes and represses them
How does alternative splicing lead to the development of male Drosophila?
- The early development TF doesn’t get produced, so theres no SXL
- When sxl gets produced later on, exon 3 is not spliced out and its contains a premature stop codon. No functional SXL is produced
- tra then gets spliced to also retain an exon with a premature stop codon, so no functional protein is produced
- DSX gets produced to have exons 1,2,3 and 5, and it will bind to and repress female sex differentiation genes
How does alternative splicing of dsx lead to vastly different functions?
The two isoforms have different DNA binding domains
How does alternative splicing contribute to the complexity of gene regulation in eukaryotes?
It allows multiple transcripts encoding proteins with multiple functions to be produced from one gene
What are the 3 possible results of alternative mRNAs?
- different isoforms with different activities
- a transcript that doesn’t get expressed even after transcription
- a transcript becoming more highly expressed after transcription
What are 4 mechanisms by which alternative mRNAs are produced from the same gene?
- Alternative promoters and TSSs
- Alternative polyadenylation sites
- RNA editing
- Alternative splicing
How does the use of alternative promoters and TSSs create alternative mRNAs?
The produced transcripts have different first exons, which changes the proteins they encode. It also changes the 5’ UTR so that sequences targeted by RNA binding proteins or miRNAs are present or absent, which changes the stability and translational efficiency of the transcript
How does the use of alternative polyadenylation sites create alternative mRNAs?
The transcripts have different C-term domains, which changes the protein’s functions. Changing the polyA tail and 3’ UTR changes protein binding site or miRNA target sequences, as well as transcript stability and translation
How does the use of RNA editing create alternative mRNAs?
If the coding sequence is altered, the polypeptide produced is changed. Changes to other sequences can change the transcript’s stability by altering protein binding or miRNA target sites
How does the use of alternative splicing create alternative mRNAs?
Polypeptides with different functions from a single gene. Inclusion or exclusion of protein binding or miRNA sites. Increase or repress gene expression
What are the 4 mechanisms of alternative splicing?
- Excluding an exon
- Retaining an intron
- Using a different 5’ or 3’ splice site
- Changes through alternative polyadenylation
