GlnB/GlnK PII proteins and regulation of the Sinorhizobium meliloti Rm1021 nitrogen stress response and symbiotic function.

Abstract

The Sinorhizobium meliloti Rm1021 Delta glnD-sm2 mutant, which is predicted to make a GlnD nitrogen sensor protein truncated at its amino terminus, fixes nitrogen in symbiosis with alfalfa, but the plants cannot use this nitrogen for growth (S. N. Yurgel and M. L. Kahn, Proc. Natl. Acad. Sci. U. S. A. 105:18958-18963, 2008). The mutant also has a generalized nitrogen stress response (NSR) defect. These results suggest a connection between GlnD, symbiotic metabolism, and the NSR, but the nature of this connection is unknown. In many bacteria, GlnD modifies the PII proteins, GlnB and GlnK, as it transduces a measurement of bacterial nitrogen status to a cellular response. We have now constructed and analyzed Rm1021 mutants missing GlnB, GlnK, or both proteins. Rm1021 Delta glnK Delta glnB was much more defective in its NSR than either single mutant, suggesting that GlnB and GlnK overlap in regulating the NSR in free-living Rm1021. The single mutants and the double mutant all formed an effective symbiosis, indicating that symbiotic nitrogen exchange could occur without the need for either GlnB or GlnK. N-terminal truncation of the GlnD protein interfered with PII protein modification in vitro, suggesting either that unmodified PII proteins were responsible for the glnD mutant's ineffective phenotype or that connecting GlnD and appropriate symbiotic behavior does not require the PII proteins.