Enhanced Symbiotic Performance by Rhizobium tropici Glycogen Synthase Mutants

Abstract

We isolated a Tn5-induced Rhizobium tropici mutant that has enhanced capacity to oxidize N,N-dimethylp-phenylendiamine (DMPD) and therefore has enhanced respiration via cytochrome oxidase. The mutant had increased levels of the cytochromes c 1 and CycM and a small increase in the amount of cytochrome aa 3 . In plant tests, the mutant increased the dry weight of Phaseolus vulgaris plants by 20 to 38% compared with the control strain, thus showing significantly enhanced symbiotic performance. The predicted product of the mutated gene is homologous to glycogen synthases from several bacteria, and the mutant lacked glycogen. The DNA sequence of the adjacent gene region revealed six genes predicted to encode products homologous to the following gene products from Escherichia coli: glycogen phosphorylase (glgP), glycogen branching enzyme (glgB), ADP glucose pyrophosphorylase (glgC), glycogen synthase (glgA), phosphoglucomutase (pgm), and glycogen debranching enzyme (glgX). All six genes are transcribed in the same direction, and analysis with lacZ gene fusions suggests that the first five genes are organized in one operon, although pgm appears to have an additional promoter; glgX is transcribed independently. Surprisingly, the glgA mutant had decreased levels of high-molecular-weight exopolysaccharide after growth on glucose, but levels were normal after growth on galactose. A deletion mutant was constructed in order to generate a nonpolar mutation in glgA. This mutant had a phenotype similar to that of the Tn5 mutant, indicating that the enhanced respiration and symbiotic nitrogen fixation and decreased exopolysaccharide were due to mutation of glgA and not to a polar effect on a downstream gene.

Metabolic Reconstruction and Modeling of Nitrogen Fixation in Rhizobium etli

“…A possible physiological explanation for the increase in nitrogen fixation rate could be that the increase of reductive power (lower NAD/NADH ratio) can be channeled to nitrogen fixation [22]. A similar effect on symbiotic nitrogen fixation is predicted for the glycogen synthase deletion, which also agrees with the observed physiological response reported for Rhizobium tropici [36]. FBA suggests that the same effect would occur for R. etli.…”

Influence of the Poly-3-Hydroxybutyrate (PHB) Granule-Associated Proteins (PhaP1 and PhaP2) on PHB Accumulation and Symbiotic Nitrogen Fixation in Sinorhizobium meliloti Rm1021

“…The concentration of PHB was determined from cells grown in YMB medium, using a spectrophotometric technique (17) as modified by Peoples and Sinskey (23). The exopolysaccharide (EPS) content was determined as previously described (20), using cultures grown in YMB for 4 days at 30°C. Glycogen was extracted from cells and determined by anthrone assay according to the procedure of Chun and Yin (6).…”

Roles of poly-3-hydroxybutyrate (PHB) and glycogen in symbiosis of Sinorhizobium meliloti with Medicago sp.

“…The roles of PHB and glycogen in rhizobia-legume symbiosis are not fully understood (Lodwig & Poole, 2003;Mandon et al, 1998;Marroqui et al, 2001;Trainer & Charles, 2006). For instance, bacteroids of determinate nodules (e.g.…”

“…A common genetic organization of the glycogen metabolism genes is found in the closely related Agrobacterium tumefaciens, Rhizobium tropici, Rhizobium leguminosarum and Mesorhizobium loti, within the glgPBCApgm operon (Lepek et al, 2002). Through analysis of a glgA mutant of R. tropici, it has been shown that there may be a link between glycogen synthase deficiency, decreased EPS, and increased symbiotic performance during symbiosis with Phaseolus vulgaris (Marroqui et al, 2001). However, the reason for the increased symbiotic efficiency of the glgA mutant is uncertain (Lodwig & Poole, 2003).…”