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MBB331_Week11-Fall-2011

Rnai of cell division gene abnormal migration sperm

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Unformatted text preview: arries out production of fresh dsRNA trigger, thus amplifying the original RNAi effect -this occurs through an siRNA-primed RdRP reaction that converts target mRNA into dsRNA, as well as possibly replicating trigger dsRNA 49 RNA interference (RNAi) in C. elegans C. • • • Caenorhabditis elegans = small nematode = soil worm soil RNAi most extensively studied in C. RNAi elegans elegans dsRNAi targets corresponding gene for dsRNAi inactivation (dominant negative interference) interference) WT WT eg. RNAi of cell division gene - abnormal migration sperm & egg nuclei 50 51 • siRNAs used as siRNAs possible way for “intracellular immunization” against viral disease disease • siRNAs used to siRNAs reduce expression of proteins for viral infection (eg. infection CCR5, the HIV-1 coreceptor) coreceptor) 52 PNAS (2003) miRNAs and gene silencing • siRNAs formed by siRNAs Dicer action on double-stranded RNAs from foreign source (transgene, virus) or transposon virus) • miRNAS are miRNAS encoded in genome and produced by cleavage of stemcleavage loop precursor RNA loop by Dicer by 53 miRNAs and gene silencing • Perfect or near Perfect perfect base-pairing between miRNA and target mRNA leads to degradation similar to RNAi, and underlies the function of the ARE destabilizing element found in unstable mRNAs unstable 54 DESTABILIZING ELEMENTS: ARE • stability of many eukaryotic mRNAs controlled by destabilizing elements in transcript • ARE: block of AU-rich sequence, 3´ untranslated region (UTR) contains AUUUA repeats • the AUUUA repeats can be placed at 3´ of stable RNA (eg. β -globin), this reduces t1/2 Lodish et al. 2000. Molecular Cell Biology 4.0. Ed. 55 DESTABILIZING ELEMENTS: ARE • instability of human ARE-containing mRNAs depends on Dicer and an miRNA (miR16) which is complementary to the ARE sequence Lodish et al. 2000. Molecular Cell Biology 4.0. Ed. 56 Hairy mRNA + 3´ Hairy UTR UTR 2. Control at level of mRNA localization Hairy Hairy mRNA, no 3´ UTR 3´ 3´ UTR analysis: inject into embryos fluorescent-labeled mRNAs, or analyze localization of lacZ-3´UTR fusions • 3´ UTRs can direct mRNA locali...
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