Interactors and effects of overexpressing YlxR/RpnM, a conserved RNA binding protein in cyanobacteria

Abstract

Throughout the tree of life RNA-binding proteins play important roles in the regulation of gene expression and RNA metabolism, but they are only poorly characterized in cyanobacteria. Here, we analyzed the predicted RNA-binding protein Ssr1238 from the model cyanobacteriumSynechocystissp. PCC 6803. Ssr1238 is encoded in a syntenic region within thenusA-ssr1238-infBoperon, similar to genes encoding such proteins in gram-positive bacteria. Overexpression of Ssr1238 for 24 h led to slightly higher levels of RNase P RNA, 44 tRNAs, and several stress-related mRNAs. Co-immunoprecipitation of proteins followed by MS analysis and sequencing of UV crosslinked, co-immunoprecipitated RNA samples identified potential interaction partners of Ssr1238. The most highly enriched transcript was RNase P RNA, and RnpA, the protein component of RNase P, was among the most highly enriched proteins, consistent with recent findings that YlxR proteins can influence the enzymatic activity of RNase P. A second highly enriched transcript derived from the 3’ region of genessl3177, which encodes a rare lipoprotein homolog, a central enzyme in cell wall remodeling during cell division. The data also showed a strong connection to the RNA maturation and modification system indicated by co-precipitation of RNA methyltransferase Sll1967, the A-adding tRNA nucleotidyltransferase Sll1253, and queuine tRNA-ribosyltransferase Slr0713, as well as riboendonuclease E and enolase. Surprisingly, cyanophycin synthetase and urease were highly enriched as well. Therefore, Ssr1238 specifically binds to two different transcripts and appears to participate in the coordination of RNA maturation, aspects of nitrogen metabolism and translation, and cell division. Our results are consistent with recent findings that theB. subtilisYlxR protein functions as an RNase P modulator (RnpM), extend its proposed role to the phylum cyanobacteria, and suggest additional functionalities.