Post-transcriptional/translational regulation (3)

Stability of mRNA can be observed by…

Comparing cell generation time and mRNA half-lives (24h vs 30mins)

Sequence associated with short half-lives

AUUUA in 3’ UTR

5’→3’ decay

1. In P-bodies

2. Decapping enzyme + XRN1 activity

3’→5’ decay

Deadenylase complex activity + exosome

Which is the predominant decay pathway?

3’→5’ decay

Structure of exosome

Barrel-like

End of exosome channel

Exonuclease activity+ endonuclease activity if exo fails

3rd decay pathway

Endonuclease activity before both 5’-3’ and 3’-5’

Why is a high iron concentration harmful?

Leads to the degradation of mononucleotides

In high iron concentration…

IRE-BP on IRE of TfR mRNA (transferrin receptor) is active = lowering of iron concentration

Blot for protein levels

Western blot

What does a lowering protein concentration + stable mRNA concentration indicate?

Translation regulation

Translational regulation in drosophila

Important role in embryo development: 2 anterior structures if not regulated

Translated ferritin

Protein that binds iron, preventing toxic levels of it

In low iron concentration…

Active IRE-BP on IRE of ferritin mRNA = no translation of ferritin

What type of molecule is lin-4?

microRNA (miRNA) that regulates gene expression post-transcriptionally

How does lin-4 regulate lin-14?

They have partial homology in the Lin-14 3’ UTR

Effect of the antisense homology/miRNA binding

Partial pairing=transcriptional repression or mRNA destabilization through de-adenylation

Which Pol transcribes miRNA?

Pol II

Primary miRNA

Folds up into dsRNA hairpin

Pre-miRNA

Forms after digestion of pri-miRNA by Drosha enzyme

Exportin for miRNA

Exportin5

Dicer action

Cleaves dsRNA into 21-23 nt fragments

Argonaute protein activity

Binds miRNA forming an RNA-induced silencing complex (RISC)

What guides the miRISC to complementary cellular RNAs?

ATP hydrolysis driving helicase activity

60% of our coding genes are thought to be…

Under miRNA-medicated control