Post-transcriptionalgenesilencing.ppt

Post-transcriptional gene silencing;;Posttranscriptional gene silencing;Other names of post-transcriptional gene silencing (PTGS) : – gene silencing – RNA silencing – RNA interference – In certain fungi: quelling RNAi can spread throughout certain organisms (C. elegans, plants). ;Short history of post-transcriptional gene silencing Definition: the ability of exogenous double-stranded RNA (dsRNA) to suppress the expression of the gene which corresponds to the dsRNA sequence. 1990 Jorgensen : Introduction of transgenes homologous to endogenous genes often resulted in plants with both genes suppressed! Called Co-suppression Resulted in degradation of the endogenous and the transgene mRNA ;1995 Guo and Kemphues: injection of either antisense or sense RNAs in the germline of C. elegans was equally effective at silencing homologous target genes 1998 Mello and Fire: -extension of above experiments, combination of sense and antisense RNA (= dsRNA) was 10 times more effective than single strand RNA;What is RNA interference /PTGS? dsRNA needs to be directed against an exon, not an intron in order to be effective homology of the dsRNA and the target gene/mRNA is required targeted mRNA is lost (degraded) after RNAi the effect is non-stoichiometric; small amounts of dsRNA can wipe out an excess of mRNA (pointing to an enzymatic mechanism) ssRNA does not work as well as dsRNA ; double-stranded RNAs are produced by: – transcription of inverted repeats – viral replication – transcription of RNA by RNA-dependent RNA- polymerases (RdRP) double-stranded RNA triggers cleavage of homologous mRNA PTGS-defective plants are more sensitive to infection by RNA viruses in RNAi defective nematodes, transposons are much more active;;;;;Dicer Double-stranded RNA triggers processed into siRNAs by enzyme RNAseIII family, specifically the Dicer family Processive enzyme - no larger intermediates. Dicer family p