post-transcriptional/translational regulation of gene expression

eukaryotic mRNA sequence motif

• AUUUA sequence in 3’ UTR

• adding AUUUA sequence to 3’ UTR of a gene that usually does not contain it dramatically destabilizes the hybrid mRNA

RNA decay

• can occur in a deadenylation-dependent or -independent manner

• decapping and 5’-3’ degradation occur most prevalently in devoted sectors of the cytoplasm called P-bodies

RNA decay in exosome

• RNA is threaded 3’-5’ into a complex of similar polypeptides that form a barrel-like structure

• exonuclease activity is present at the end of the channel

• endonuclease activity is present near the exit in case exonuclease activity fails

endonuclease-meditated RNA decay

RNA is first cleaved in the middle and then degraded simultaneously by both exonucleolytic degradation pathways

mammalian transferrin receptor (TfR)

needed for the import of iron into the cell

stability of mammalian transferrin receptor (TfR) mRNA

regulated by intracellular levels of iron

• high iron: no iron response element binding protein present, mRNA is degraded

• low iron: active iron response element binding protein, little mRNA degradation

translational regulation

• abundance of mRNA usually reflects protein levels (more mRNA = more protein)

• when relationship appears skewed, may indicate that protein synthesis or stability of protein is regulated

inhibition of mRNA translation in Drosophila embryo

• hunchback mRNA is distributed uniformly throughout the fertilized egg, but the protein is present in a very steep anterior to posterior gradient

• in mutants, nanos protein is absent and hunchback protein is present uniformly in early embryo, preventing proper larva development and leading to death

ferritin

intracellular protein that binds iron ions, thereby precenting the accumulation of toxic levels of free iron ions

regulation of stability of ferritin mRNA

dependent on iron levels

• high iron: IRE-BP absent, mRNA is translated

• low iron: IRE-BP can bind to IREs in 5’ UTR of ferritin mRNA, inhibiting translation

developmental timing mutants in C. elegans

• lin-4 lf mutants look very similar to lin-14 gf mutants

• lin-4 encodes a small RNA that has considerable homology to regions of the lin-14 3’ UTR

• lin-4 RNA is synthesized as a longer precursor then it has to be processed, after which it binds its target sites and affects translation of the lin-14 mRNA

microRNA (miRNA) biogenesis

1. primary miRNA transcript (pri-miRNA) folds into dsRNA hairpin

2. folded miRNA is digested by an enzyme called Drosha to generate pre-miRNA

3. Exportin5 takes pre-miRNA into the cytoplasm via the NPC

4. dicer cleaves dsRNA into 21-23nt fragments that are then bound by an argonaute protein in an RNA-induced silencing complex, forming a miRISC

5. ATP hydrolysis drives the unwinding activity of miRISC, which then uses the miRNA product as a guide to target the complex to complementary cellular RNAs

6. miRNA binding to their target mRNAs usually occurs through interactions in the 3’ UTR and the pairing is usually imperfect

dicer

RNAase III like enzyme

result of miRNA and mRNA binding

block translation or destabilize the mRNA target through de-adenylation

regulatory roles of miRNA

• metabolism

• tissue growth

• neural development

• developmental timing

• stem cell biology/pluripotency

• cancer