RNA interference (RNAi) (lect 4)

Question 1

miRNA

Answer 1

-21-25nts encoded by endogenous genes
-hairpin precursors
-recognise multiple targets (regulate other genes)

Question 2

siRNA

Answer 2

-21-25nts encoded by exogenous genes
-ds precursors
-recognise one target

Question 3

Processing of miRNA

Answer 3

-pri-miRNA transcribed
-microprocessor protein complex (Drosha and DGCR8) remove flanking seqs to make pre-miRNA (just hairpin structure) (DGCR8 guides Drosha to correct seq and Drosha acts as a RNase and cleaves)
-exported to cytoplasm by exportin 5 and Ran (RanGAP, its GTPase, triggers disassembly in cytoplasm, and then exportin 5 and Ran are recycled back into nuc)
-Dicer matures it to miRNA (driven by ATP hydrolysis)
-one strand of miRNA selected

Question 4

What is RNA interference (RNAi)?

Answer 4

biological response to dsRNA
-regulates expression of genes (sequence-specific silencing)
-mediates resistance to endogenous foreign (eg. viral) and exogenous pathogenic nucleic acids

Question 5

RISC complex in RNAi

Answer 5

RISC = RNA-induced silencing complex

-during RISC assembly, mRNA is unwound and then RISC can bind to single strand
-RISC either inhibits translation or triggers mRNA cleavage and degradation

Question 6

How is RNAi used in research?

Answer 6

-siRNAs can be added to degrade naturally-occurring cellular RNA
-reduces protein levels to study function of targeted protein