Mechanistic studies of a new family of bacterial RNA chaperones

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Small non-coding RNAs (sRNAs) regulate gene expression in essentially all bacterial species. My laboratory showed that the regulation of gene expression in the F family of bacterial plasmids is controlled by an RNA chaperone, called FinO, that facilitates the interactions of antisense RNAs with their mRNA targets. In a new collaboration with Xavier Charpentier (Lyons), we have now shown that FinO-like chaperones exist in a variety of bacterial species where they control diverse regulatory processes. Charpentier has recently used genomic approaches to identify an sRNA-mRNA pair that regulates DNA transduction in L. pneumophila and he has shown that L. pneumophila FinO is required for this regulatory network. Building on this, we will work to uncover the mechanism by which L. pneumophila FinO regulates sRNA function. My lab will utilize our expertise in the biochemical, biophysical, and crystallographic study of protein-nucleic acid interactions to derive structural mechanisms for the action of L. pneumophila FinO. In preliminary work, we have already shown that L. pneumophila FinO specifically binds its sRNA and we have isolated and crystallized a minimal RNA-binding domain of L. pneumophila FinO. We hypothesize that L. pneumophila FinO acts by destabilizing hairpin structures with the bound RNA that inhibit base pairing between the sRNA and the mRNA target. We will test this hypothesis using a set of biochemical assays and will further probe the structural principles underlying the interactions between L. pneumophila FinO and RNA through crystallographic and small angle X-ray scattering approaches. Our mechanistic models will be tested through analysis of the effects of structure-guided site directed mutants in living cells in collaboration with the Charpentier lab. This will set the basis for the comparison of the function of the FinO family of proteins across diverse bacterial species. This program will offer outstanding training opportunities for students to gain experience in cutting edge structural biology within my lab, as well as an opportunity to extend their training to areas of microbiology and genomics through our international collaborations.**