A Family of Small Intrinsically Disordered Proteins Involved in Flagellum-Dependent Motility in Salmonella enterica.

By screening a collection of mutants deleted for genes encoding small proteins of ≤60 amino acids, we identified three paralogous small genes (, , and ) required for wild-type flagellum-dependent swimming and swarming motility. The , , and genes encode small proteins of 55, 60, and 60 amino acid residues, respectively. A bioinformatics analysis predicted that these small proteins are intrinsically disordered proteins, and circular dichroism analysis of purified recombinant proteins confirmed that all three proteins are unstructured in solution. A mutant deleted for STM14_1829 showed the most severe motility defect, indicating that among the three paralogs, STM14_1829 is a key protein required for wild-type motility. We determined that relative to the wild type, the expression of the flagellin protein FliC is lower in the Δ_ mutant due to the downregulation of the operon encoding the FlhDC master regulator. By comparing the gene expression profiles between the wild-type and Δ_ strains via RNA sequencing, we found that the gene encoding the response regulator PhoP is upregulated in the Δ_ mutant, suggesting the indirect repression of the operon by the activated PhoP. Homologs of STM14_1829 are conserved in a wide range of bacteria, including and We showed that the inactivation of STM14_1829 homologs in and also alters motility, suggesting that this family of small intrinsically disordered proteins may play a role in the cellular pathway(s) that affects motility. This study reports the identification of a novel family of small intrinsically disordered proteins that are conserved in a wide range of flagellated and nonflagellated bacteria. Although this study identifies the role of these small proteins in the scope of flagellum-dependent motility in , they likely play larger roles in a more conserved cellular pathway(s) that indirectly affects flagellum expression in the case of motile bacteria. Small intrinsically disordered proteins have not been well characterized in prokaryotes, and the results of our study provide a basis for their detailed functional characterization.