Identification and characterization of novel RNA-binding proteins in Salmonella Typhimurium

RNA binding proteins (RBPs) are important effectors of post-transcriptional control, affecting the stability, maturation, translation, or localization of bound RNAs. They thereby exert a crucial role in bacterial physiology, pathogenicity and stress. Although some bacterial RBPs are well-characterized, discovery of novel RBPs in bacteria has been largely serendipitous. Only recently global approaches for RNA-binding proteome (RBPome) capture in prokaryotes have been developed, revealing many previously unidentified RNA-interacting proteins. The increasing number and diversity of potential RBPs further adds to the complexity of the bacterial RBPome and underscores the importance of identifying and characterizing RBPs.
Our group recently developed an approach termed CoCAP to globally capture RBPs in bacteria. Applying this method to the enteropathogenic proteobacteria Salmonella Typhimurium and Campylobacter jejuni, we were able to capture known RBPs, such as RNA chaperones Hfq and ProQ (in Salmonella) as well as RNases, translation factors and ribosomal proteins. Importantly, we also identified a considerable number of novel RBP candidates, many of which lack known RNA binding domains and have well-established cellular functions unrelated to RNA. These include diverse processes including cell cycle control, metabolism, replication, and virulence. We speculate that such unconventional RBPs may moonlight as RNA regulators in addition to their canonical roles in response to distinct conditions, or bound RNAs act as riboregulators and impact on the proteins' function, stability or localization. To ensure that the identified candidates are bona fide RBPs, we have been validating selected candidates by applying diverse techniques. We now aim to further characterize their interaction with RNA and elucidate the regulatory consequences of these interactions. Expanding our novel RBP-capture approach to a variety of infection- and stress-relevant conditions will further broaden our knowledge of the RBP repertoire of bacteria and aid our understanding of post-transcriptional regulation and its effects on bacterial physiology.