The Sigma Factor AlgU Regulates Exopolysaccharide Production and Nitrogen-Fixing Biofilm Formation by Directly Activating the Transcription of in A1501.

A1501, a plant-associated diazotrophic bacterium, prefers to conform to a nitrogen-fixing biofilm state under nitrogen-deficient conditions. The extracytoplasmic function (ECF) sigma factor AlgU is reported to play key roles in exopolysaccharide (EPS) production and biofilm formation in the genus; however, the function of AlgU in A1501 is still unclear. In this work, we mainly investigated the role of in EPS production, biofilm formation and nitrogenase activity in A1501. The mutant Δ showed a dramatic decrease both in the EPS production and the biofilm formation capabilities. In addition, the biofilm-based nitrogenase activity was reduced by 81.4% in the Δ mutant. The transcriptional level of , a key Psl-like (a major EPS in A1501) synthesis-related gene, was almost completely inhibited in the mutant and was upregulated by 2.8-fold in the -overexpressing strain. A predicted AlgU-binding site was identified in the promoter region of . The DNase I footprinting assays indicated that AlgU could directly bind to the promoter, and β-galactosidase activity analysis further revealed mutations of the AlgU-binding boxes drastically reduced the transcriptional activity of the promoter; moreover, we also demonstrated that AlgU was positively regulated by RpoN at the transcriptional level and negatively regulated by the RNA-binding protein RsmA at the posttranscriptional level. Taken together, these data suggest that AlgU promotes EPS production and nitrogen-fixing biofilm formation by directly activating the transcription of , and the expression of AlgU is controlled by RpoN and RsmA at different regulatory levels.