A NEW GENETIC-LOCUS IN SINORHIZOBIUM-MELILOTI IS INVOLVED IN STACHYDRINE UTILIZATION

Stachydrine, a betaine released by germinating alfalfa seeds, function s as an inducer of nodulation genes, a catabolite, and an osmoprotecta nt in Sinorhizobium meliloti. Two stachydrine-inducible genes were fou nd in S. muliloti 1021 by mutation with a Tn5-luxAB promoter probe. Bo th mutant strains (S10 and S11) formed effective alfalfa root nodules, but neither grew on stachydrine as the sole carbon and nitrogen sourc e. When grown in the absence or presence of salt stress, S10 and S11 t ook up [C-14]stachydrine as well as wild-type cells did, but neither u sed stachydrine effectively as an osmoprotectant. In the absence of sa lt stress, both S10 and S11 took up less [C-14]proline than wild-type cells did. S10 and S11 appeared to colonize alfalfa roots normally in single-strain tests, but when mixed with the wild-type strain, their r hizosphere counts were reduced more than 50% (P less than or equal to 0.01) relative to the wild type. These results suggest that stachydrin e catabolism contributes to root colonization. DNA sequence analysis i dentified the mutated locus in S11 as putA, and the luxAB fusion in th at gene was induced by proline as web as stachydrine. DNA that restore d the capacity of mutant S10 to catabolize stachydrine contained a new open reading frame, stcD. All data are consistent with the concept th at stcD codes for ari enzyme that produces proline by demethylation of N-methylproline, a degradation product of stachydrine.