pre-mRNA processing- Lecture 25 Flashcards
What is the post-transcriptional process of pre-mRNA processing?
capping, splicing, and polyadenylation
What is a 5’ cap composed of?
guanosine nucleotide that has been modified to contain a methyl group at the 7 position of the purine ring structure linked to the 5’ end of the transcription in an unusual 5’ to 5’ phosphodiester bond
What is the function of a 5’ cap?
prevents degridation of mRNA/pre-mRNA by exonucleases (which recognize 5’ ends) and is a recognition site for binding of proteins that recruit a ribosome
Introns contain a ____ sequence at 5’ splice site, a _____ near the 3’ splice site, and the ____ sequence at the 3’ splice site.
GU
branch point (normally an A residue in the context of a loose consensus sequence)
AG
What catalyzes the splicing reaction?
splicesomes (large complex of nearly 200 proteins and several snRNPs)
What happens during the splicing process?
two transesterification reactions
- 2’ OH of the branch point (A) attacks the 5’ splice site between the exon and the GU sequence (resulting in a circular bond between the branch point and the GU)
- OH now on the end of the exon attacks the 3’ branch site of the intron between the AG and the next exon (resulting in the binding of the two exons and a Lariat RNA)
How do splisosomes know when they have reached the proper 5’ GU and 3’ AG?
ESE sequences in exons bind SR proteins which indicate to the splicing machinery when they are in close proximity of the proper sequence
What is alternative splicing and why is it important?
the selective splicing of exons/introns in different arrangements that increases the number of possible mRNA products
What are the forms of alternative splicing?
alternative last exon, mutually exclusive, alternative 5’ splice site, alternative first exon, cassette exon, alternative 3’ splice site, and intron retention
How is alternative splicing achieved?
by ESE and ESS sequences that are bound by different SR proteins which regulate transcription
It is estimated that ~_____% of all point mutations that cause genetic disease are due to changes in splicing caused by these mutations.
15
When would a silent mutation be harmful?
when the changed sequence is part of a cis regulator (eg. ESE or ESS)
Where is the polyadenylation signal located and what is it?
at the end of the 3’ UTR
most often the sequence is AAUAAA which indicates that endonucleolytic cleavage is necessary 10-30 nucleotides down stream
How is polyadenylation performed?
the free 3’ hydrozyl is acted upon by poly(A) polymerase which adds 100-200 A residues in a template-independent fashion and bound by PABP
