Comparative genomics of Sinorhizobium meliloti isolates differed from salt tolerance

Секция 2. Генетика и селекция растений, животных и микроорганизмов Подсекция 2.3. Генетика микроорганизмо

[Identification of Sinorhizobium meliloti genes influencing synthesis of surface polysaccharides and competitiveness]

Onishchuk OP, Sharypova LA, Kurchak ON, Becker A, Simarov BV. [Identification of Sinorhizobium meliloti genes influencing synthesis of surface polysaccharides and competitiveness]. Genetika. 2005;41(12):1337-1342.A new approach to isolating mutants with altered composition of capsular polysaccharides (CPS) and lipopolysaccharides (LPS) in nodule bacteria of alfalfa Sinorhizobium meliloti based on analysis of their respiratory activity was proposed. Random Tn5-mob mutants of symbiotically effective strain SKhM1-105 were tested for slime-production ability and coloration on diagnostic media containing the indicator of reducing equivalents, triphenyltetrasolium chloride (TTC), the inhibitor of respiratory activity, 2-methyl-4-chlorphenoxybutyric acid and Congo Red, the stain for LPS and exopolysaccharides (EPS). Electrophoretic analysis (SDS-PAGE) of polysaccharides in seven mutants, markedly differing from the parental strain with respect to their growth on diagnostic media demonstrated that (1) the production of CPS was either decreased (in mutants T64 and T835) or blocked (in T71, T94, T124, T134, and T170); (2) the R form of LPS had changed mobility (in T134); (3) the S form of LPS contained only one component instead of two (T71). In symbiosis with alfalfa Medicago sativa, all mutants exhibited a decreased competitive ability for adsorption on roots of host alfalfa plant, compared to the parental strain. At early stages of symbiosis, mutants had a tendency toward a decrease in the number of nodules, in comparison with the parental strain (a statistically significant decrease was observed in mutants T71 and T64). In mutants T64, T71, and T134, nodulation competitiveness was lower than in the parental strain. Analysis of Tn5-mob tagged sequences of the mutants revealed homologies with the genes encoding methionine synthase, luciferase-like monooxygenase, UDP-glucuronic acid epimerase, sensor hystidine kinase, acetoacetyl-CoA-reductase, oligopeptide uptake ABC transporter, and a transcription activator

The eff-482 locus of Sinorhizobium meliloti CXM1-105 that influences symbiotic effectiveness consists of three genes encoding an endoglycanase, a transcriptional regulator and an adenylate cyclase

Sharypova LA, Yurgel SN, Keller M, Simarov BV, Pühler A, Becker A. The eff-482 locus of Sinorhizobium meliloti CXM1-105 that influences symbiotic effectiveness consists of three genes encoding an endoglycanase, a transcriptional regulator and an adenylate cyclase. MOLECULAR AND GENERAL GENETICS. 1999;261(6):1032-1044.The mutant T482 of Sinorhizobium meliloti CXM1-105, which carries a Tn5 insertion on megaplasmid 1, exhibits an enhanced symbiotic efficiency phenotype. Three genes, eglC, cya3 and syrB2, were identified in the eff-482 region tagged by the Tn5 insertion in T482. The eglC gene encodes an endoglycanase which contributes to the depolymerization of the exopolysaccharide succinoglycan. The N-terminal region of the predicted cya3 gene product was similar to eukaryotic-type adenylate cyclases from Brevibacterium liquefaciens and Streptomyces coelicolor. Four contiguous tetratricopeptide repeats which are known to mediate protein-protein interactions were identified in the C-terminal portion of Cya3. Complementation analysis demonstrated that cya3 indeed encodes a functional adenylate cyclase. A central helix-turn-helix DNA-binding motif and a putative C-terminal coiled-coil structure implicated in protein oligomerization were found in SyrB2. Extra copies of the syrB2 gene negatively affect transcription of both syrB2 itself and cya3. The Tn5 insertion in T482 was localized between the divergently transcribed genes eglC and syrB2. It eliminated eglC function and slightly stimulated transcription of both syrB2 and cya3, which lies downstream of syrB2. Mutants carrying insertions of the lacZ-Gm interposon in the genes eglC, syrB2 and cya3 exhibit the same phenotype as mutant T482, indicating that these three genes influence symbiotic efficiency