The conserved noncoding RNA ModT coordinates growth and virulence in Clostridioides difficile.

Bacterial noncoding RNAs fulfill a variety of cellular functions as catalysts, as scaffolds in protein complexes or as regulators of gene expression. They often exhibit complex tertiary structures that are a key determinant of their biochemical function. Here, we characterize the structured "raiA motif" RNA from Clostridioides difficile, which is conserved in more than 2,500 bacterial species from the phyla Bacillota and Actinomycetota. We show that its transcript abundance and stability in exponentially growing bacteria rivals that of ribosomal RNAs. Deletion of the "raiA motif" RNA is associated with delayed transition into stationary phase, and changes in stationary phase pathways such as spore formation, hence we rename it ModT (modulator of transition phase). Mechanistically, we show that ModT-mediated changes in cellular cyclic di-GMP levels are linked to the pronounced sporulation defect in the modT mutant. Importantly, we show that expression profiles and isoform patterns of ModT are conserved in Clostridium perfringens and Paeniclostridium sordellii, and that these orthologs can functionally complement ModT in C. difficile. Chemical structure probing of ModT in vivo reveals dynamic refolding and provides initial evidence for a potential association of ModT with proteins. In summary, our findings indicate that ModT fulfills a conserved role in regulating growth transitions in bacteria and provide a crucial step towards delineating its molecular mechanism.