Topic 8: Posttranscriptional Gene Regulation

Where is newly transcribed precursor mRNA (pre-mRNA) processed to translatable mature mRNA in eukaryotes?

Nucleus

5’-UTRs and 3’-UTRs are part of what?

The first and last exon, and are untranslated regions

What do UTRs do?

Regulate mRNA stability and translational initiation

When is the 5’-CAP and 3’ poly-A tail added?

5’-CAP is added early in transcription and 3’ poly-A tail is added soon after cleavage of mRNA by CPSF

What does pre-mRNA contain?

Alternating coding segments and UTRs (exons) and non-coding segments (introns)

What does the removal of introns by splicing during transcription do?

Generates the open reading frame (ORF) consisting of a continuous stretch of codons (flanked by start and stop codons) and the UTRs

What is RNA processing coordinated by?

Recruitment of enzymes to CTD of RNA pol II

Once processed, where is mRNA transported?

Exported from nucleus into cytoplasm to associate with ribosomes (for translation)

What does the 5’-CAP contain?

7-methylguanosine (7-MeG) is added to 5’ end of pre-mRNA via 5’,5’ triphosphate linkage by guanylyltransferase after the first 20-30 nt of the transcript have been added

What does 5’-CAP do?

• Protects mRNA from degradation by 5’-3’ exoribonucleases because a 5’ phosphate has been replaced with a methylated guanosine (steric hindrance)

• Also acts as a ribosome binding site whereby the 5’ cap recruits the cap binding complex (CBC)

What does the CBC do?

Helps recruit a small ribosomal subunit (40S) to the mRNA which then scans for the start codon (and to the right end)

What did the classical studies of 5’ cap do?

Synthesized 5’-capped and uncapped mRNA by transcription assay using wheat cell extracts with labels labeled GTP and SAM (produces 7-meG)

• sucrose density gradient centrifugation showed that only 5’-capped mRNAs were bound to ribosomes