CMB2001/L10 Post-transcriptional Control IIb Flashcards
Using casein mRNA, explain how mRNA levels are regulated.
Expressed in mammary gland
mRNA increases 70-fold on stimulation by prolactin
Transcription increases 2-fold as half-life increases 40-fold
PolyA tail length increased
3’ UTR of RNA binds proteins which aid in stabilisation
Give 3 reasons for mRNA degradation.
Damaged mRNA
Incorrectly transcribed/processed mRNA
Control gene expression
What must occur to mRNA before exonucleases can gain access?
Closed loop must be broken
Describe deadenylation-dependent decay.
Decapping of mRNA by DCP1&2 and 5’-3’ degradation by exonucleases
PolyA tail removal by CCR4-NOT or PARN by removing adenine residues (then exosome complex degrades 3’-5’)
Give 2 decapping enzymes.
DCP1
DCP1
Give 2 endonucleases.
Argonaute
Swt1
Smg6
Give a deadenylase enzyme.
CCR/Not complex
Which enzyme degrade mRNA:
a) 5’-3’
b) 3’-5’?
a) XRN1
b) the exosome
Give 2 nucleases in the exosome.
RRP6
RRP44
Give another use of the exosome.
RNA turnover and processing
Give 2 other functions of XRN1.
Involved in RNA turnover and processing
Involved in transcription termination
Functions after decapping of mRNA
Describe nonsense mediated decay (NMD).
Mistakes in RNA are detected and mRNA targeted for degradation
Give 3 ways in which premature stop codons can occur in mRNA.
Errors in:
Transcription
Splicing
Editing
Polyadenylation
Mutations
Describe nonsense-mediated decay as a surveillence mechanism.
Targets mRNA with premature stop codons (nonsense)
Recognises presence of stop codons and tags mRNA for degradation
Stops production of truncated, potentially harmful proteins
Describe the role of the exon junction complex (EJC).
Deposited near splice junctions during mRNA splicing
Marks spliced mRNAs
If ribosome encounters premature stop codon and EJC remains downstream, indicated mRNA is defective
What do these abbreviations mean:
a) RNAi
b) siRNA
c) miRNA
d) RISC?
a) RNA interference
b) small inhibitory RNA
c) micro RNA
d) RNA-induced silencing complex
Describe siRNA.
21-23 nucleotide RNAs
Perfect complementary to target RNA
Thought to be mainly viral defence mechanism
Leads to degradation of target RNA
Describe miRNA.
21-23 nucleotide RNAs
Imperfect complementary to target RNA
Key gene regulatory mechanism in the cell
Leads to block in translation
Describe siRNA production.
dsRNA precursor
Dicer enzyme recognises and processes long dsDNA into fragments
Incorporation into RISC
Target recognition and cleavage
Describe miRNA production.
Primary miRNA (pri-miRNA)
Drosha enzyme with cofactor DGCR8 processes pri-mRNA into shorter precursor
Export to cytoplasm
Dicer enzyme processes into mature miRNA duplexes
One strand incorporation into RISC - other is degraded
Target recognition and regulation
What happens to UTRs during embryonic development?
Get longer
How does the length of mRNA differ in proliferating cells?
They’re shorter
Give 3 uses of siRNA in research.
Gene knockdown
Functional genomics
Disease modelling
Target validation
Therapeutic applications
Pathway analysis
High-throughput screening