S-Nitrosoglutathione cytotoxicity toMycobacterium smegmatisand its use to isolate stationary phase survival mutants

Abstract

We report that stationary phase Mycobacterium smegmatis is more sensitive than exponential phase cells to the nitric oxide donor S-Nitrosoglutathione (GSNO). This finding was used to select for both spontaneous and transposon mutants of M. smegmatis with increased resistance to GSNO in stationary phase. Some of these mutants were also defective in stationary phase survival, demonstrating a link between sensitivity to GSNO and stationary phase survival. Transduction of the disrupted region from seven selected mutants indicated that the transposon insertion was linked to the GSNO-resistance and stationary phase survival phenotypes. For five mutants, the disrupted genes were identified. Three were homologous to genes with possible roles in nutrient scavenging, including: (i) a putative amino acid efflux pump, (ii) a putative thioesterase and (iii) an enoyl-CoA-hydratase. One mutant was disrupted in the atpD gene, encoding the beta chain of F1 F0 ATP synthase. We independently isolated a stationary phase survival mutant disrupted in the atpA gene (encoding the alpha chain) of the F1 F0 ATP synthase of the same operon, suggesting an important role for efficient ATP synthesis in stationary phase survival.

Starvation/stationary-phase survival of Rhodococcus erythropolis SQ1: a physiological and genetic analysis

“…Similar genes have been identified in stationary/starvation deficient mutants of other organisms. Some of those genes are involved in amino acid metabolism, such as hprT (hypoxanthine-guanine phosphoribosyl transferase) in Staphylococcus aureus (Watson et al, 1998a), lysE (exporter of lysine and arginine) in Mycobacterium smegmatis (Smeulders et al, 2004) and ilvE (branched-chain amino acid aminotransferase) in Sinorhizobium meliloti (Uhde et al, 1997). Others are cytochrome-associated, e.g.…”