An Azorhizobium caulinodans ORS571 mutant with deletion of a gene encoding a TIGR02302 family protein overproduces exopolysaccharides and is defective in infection into plant host cells

Abstract

ABSTRACTAzorhizobium caulinodans is a microsymbiont of Sesbania rostrata Bremek. & Oberm., and is able to fix nitrogen in both the free-living and symbiotic states. In this study, we focused on the ggm gene (locus tag, AZC_4606) that encodes a putative membrane protein belonging to the TIGR02302 family. Although the genes encoding TIGR02302 family protein are distributed in a wide range of alpha-proteobacteria including rhizobia, the functions of this protein are still unknown. To investigate the functions of this protein in A. caulinodans, we made a ggm mutant, and analyzed its phenotypes. The ggm mutant produced more bubbles than the wild-type strain in L3 + N medium liquid cultures, and formed mucoid colonies on L3 + N medium agar plates, suggesting that the ggm mutant overproduced exopolysaccharides (EPSs). The amounts of EPSs produced by the ggm mutant on L3 + N plates were about 1.3-fold higher than those by the wild-type strain, and expression levels of EPS production-related genes in the ggm mutant grown in L3 + N liquid medium were about 2- to 4-fold higher than those of the wild-type strain. In addition, the stem nodules formed by the ggm mutant on the stems of S. rostrata showed little or no nitrogen-fixing activity. By microscopic analyses, large infection pockets and a few infected cells were observed in the stem nodules formed by ggm mutant, suggesting that the ggm mutant is defective in invasion into plant cells. Taken together, our results suggest that Ggm is involved in EPS production and that adequate levels of EPS production are required for A. caulinodans to invade into host cells.