Insertion of a Divergent GAF-like Domain Defines a Novel Family of YcgR Homologues That Bind c‑di-GMP in Leptospirales

The Leptospiraceae family, which includes the genera Leptospira, Leptonema, and Turneriella, is an ecologically diverse group that includes saprophytic strains from soil and water as well as important pathogenic strains. Adaptation to these multiple environments relies strongly on signal transduction to adjust their morphology, motility, and metabolism to the changing environmental conditions. Members of the genus Leptospira distinguish themselves among spirochetes for having an elevated number of signal transduction genes. In this study, we describe a novel signal transduction protein that has gained multiple paralogues in the Leptospiraceae. These proteins are members of the YcgR/DgrA/MotI family, whose orthologs in several bacterial lineages have been shown to regulate the flagellar motor upon binding to c-di-GMP through their N-terminal PilZ domain. Unlike previously described versions of YcgR, the spirochetal proteins are characterized by the insertion of a divergent GAF domain within their N-terminal PilZ domain. We show that one member of this protein family from Leptospira interrogans is still a monomeric c-di-GMP binding protein and that these novel YcgR-like proteins have mostly replaced other members of the YcgR family in Leptospiraceae. Marked divergence among the paralogs suggests this family’s expansion was accompanied by neofunctionalization, with the likely emergence of novel interactions in the signal transduction network controlling the flagellum rotor and other processes affected by changes in levels of c-di-GMP.