Les courts ARN chez C. elegans : spécificité et fonction des protéines argonautes

Authors: Boisvert, Marie-Ève
Advisor: Simard, Martin
Abstract: The molecular characteristics of the RNAi and microRNA pathways are different. In the RNAi pathway, fully base-paired dsRNA molecules trigger the production of small interfering RNAs (siRNAs), which lead to the degradation of the complementary targeted mRNA. On the other hand, the stem-looped miRNA precursor is processed in mature miRNA, which then imperfectly interacts with the mRNA target, leading to the blocking of its translation. In the worm C. elegans, each of the RNAi and miRNA pathways needs its specific Argonautes proteins. RNAi requires RDE-1, while ALG-1 and ALG-2 act in the miRNA pathway. The restriction of siRNAs and miRNAs to specific Argonaute proteins might reflect the recognition of the trigger by specific factors targeting it to the correct Argonaute protein. To better understand the importance of the trigger in the selection of the adequate Argonaute and pathway, we designed a dsRNA trigger containing both miRNA and siRNA sequences. This chimeric molecule can rescue successfully the loss of function of the miRNA let-7, and can also initiate the gfp gene silencing by RNAi. We demonstrated that RDE-1 and the dsRNA-binding protein RDE-4 are essential for RNAi induced with our trigger, but are not involved in the let-7 function of the chimera molecule. On the other hand, we showed that ALG-2 is strictly required for the miRNA function, but not for the RNAi function. Interestingly, we also found that the let-7 miRNA processed from our molecule has a limited lifetime, while the RNAi response, initiated from the same dsRNA, is maintained for a longer period. We suggest that the specificity of the Argonautes and thus the choice of the small RNA pathway is not determined by the type of RNA trigger, but rather by their respective molecular response. Furthermore, we also tried to understand the roles of ALG-1 and ALG-2. An immunoprecipitation of these proteins and an analysis of the protein and RNA interactors were carried out.
Document Type: Mémoire de maîtrise
Issue Date: 2007
Open Access Date: 13 April 2018
Permalink: http://hdl.handle.net/20.500.11794/19666
Grantor: Université Laval
Collection:Thèses et mémoires

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