lecture 7 Flashcards
What are the main RNA processing events in pre-mRNA?
Capping
Splicing (including alternative splicing)
Polyadenylation
RNA Editing
How are RNA processing events coupled to transcription?
Through the RNA polymerase II (Pol II) CTD, which acts as a landing pad for processing factors.
What is the structure and function of the 5’ m7G cap?
Structure: Unusual, with both sugar-phosphate bonds and base methylation.
Functions:
Protects mRNA from degradation by 5’-3’ nucleases.
Facilitates splicing and nuclear export.
Critical for translation via protein binding to CBP80/CBP20 (nucleus) and eIF4 complex (cytoplasm).
: What are the steps in the capping process?
Addition of GpppN structure.
Methylation of the cap, which alters the chemical behaviour of the base.
What are the conserved sequences involved in splicing?
5’ splice site: AGGURAGU.
3’ splice site: YYYYYYNCAGGU.
Branch site: YURAC.
What is the mechanism of splicing?
Step 1: 5’ splice site is cut, forming a bond between the 5’ end of the intron and the branch site.
Step 2: 3’ splice site is cut, releasing the intron as a lariat and joining the exons.
What is the spliceosome?
A large complex with >200 proteins and several snRNPs (U1, U2, U4, U5, U6) that catalyse intron removal. It includes RNA-binding proteins, ATPases, and GTPases.
How does alternative splicing expand the proteome?
It generates multiple protein isoforms from a single gene through mechanisms like:
Exon skipping.
Intron retention.
Alternative splice sites (5’ and 3’).
Give an example of extensive alternative splicing.
The Dscam gene in Drosophila, which generates 38,016 isoforms.
What diseases are linked to splicing defects?
Spinal muscular atrophy (infant mortality).
Retinitis pigmentosa (blindness).
Myotonic dystrophy (muscle wasting).
What is polyadenylation and its significance?
Adds a poly(A) tail (~250 nucleotides) to the 3’ end of mRNA.
Functions:
Enhances RNA export.
Stabilises the mRNA.
Promotes translation.
Provides a binding site for poly(A)-binding proteins.
What sequences are required for polyadenylation?
Conserved AAUAAA sequence (10-35 nucleotides upstream of the site).
G/U-rich tract downstream of the site.
USE (U-rich upstream element).
What are the types of RNA editing?
Insertion/Deletion of nucleotides.
Modification:
A-to-I (Inosine recognised as G).
C-to-U or U-to-C.
What are the effects of RNA editing?
Alters the coding sequence.
Changes mRNA properties.
Impacts RNA binding and translation.
How does editing affect diseases and development?
Diseases: Atherosclerosis, cancer.
Development: Brain function and Drosophila development.
Parasites: Trypanosoma and Leishmania.
How do introns and exons vary in length?
Introns are generally longer than exons in mammals, ranging from 80 to 10,000 nucleotides.
The dystrophin gene has one intron over 210,000 nucleotides.
Which gene has the most introns, and what is its significance?
Dystrophin: >50 introns, linked to Duchenne muscular dystrophy.
What are key steps of regulation in gene expression?
Transcription control.
RNA processing (capping, splicing, polyadenylation, editing).
Translation control.
Protein activity control.
Why is mRNA localisation important?
Ensures localised protein synthesis.
Prevents expression in incorrect locations (e.g., myelin basic protein).
Enhances translation efficiency at specific sites.
