[
International Worm Meeting,
2017]
MicroRNAs are small non-coding RNA who regulates the expression of several genes. The miRNA pathway effector complex, the miRISC (miRNA induced silencing complex), is formed by the association of an argonaute protein with a miRNA. The subsequent binding of the miRISC to its mRNA target can either repress translation or degrade the mRNA. To further understand the miRNA gene regulation mechanisms, it is necessary to characterize new important molecular partners involved in the miRNA silencing. Using different genetic and molecular screens, we identified new miRISC interactors. Among them, we found a DnaJ chaperone protein as an interactor of a C. elegans miRNA-specific Argonaute ALG-1 in the yeast two-hybrid system. Different genetic assays indicate that the DnaJ chaperone is important for the microRNA pathway in animals. Interestingly, observations made with Drosophila cells extract suggested that Heat Shock Proteins or HSPs are required for small RNA loading onto Argonaute proteins (Iwasaki and al, Molecular Cell, 2010). It is also known that DnaJ chaperone proteins can provide specificity to HSPs in order to guide them to specific proteins. We therefore hypothesize that the DnaJ chaperone interacting with ALG-1 recruits HSPs proteins in order to permit loading of miRNAs on this Argonaute. To test this model, we will first demonstrate that DnaJ protein interacts with ALG-1 associated to the loading complex in vivo with the newly generated animals expressing endogenously HA-tagged DNaJ gene. We will next test whether DnaJ is required for: 1) the interaction of HSPs with ALG-1 and; 2) the loading of miRNA onto ALG-1 using both RNAi and KO animals. Taken together, this study will provide mechanistic insights on how microRNA (and potentially others small RNAs in worms) are loaded specifically onto the proper Argonaute proteins.