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SummaryMetal ion homeostasis is regulated principally by metalloregulatory proteins that control metal ion uptake, storage and efflux genes. We have used transcriptional profiling to survey Bacillus subtilis for genes that are rapidly induced by exposure to high levels of

We examined the effects of nitric oxide (NO) and sodium nitroprusside (SNP) on Bacillus subtilis physiology and gene expression. In aerobically growing cultures, cell death was most pronounced when NO gas was added incrementally rather than as a single bolus, suggesting that the length of exposure was important in determining cell survival. DNA microarrays, Northern hybridizations, and RNA slot blot analyses were employed to characterize the global transcriptional response of B. subtilis to NO and SNP. Under both aerobic and anaerobic conditions the gene most highly induced by NO was hmp, a flavohemoglobin known to protect bacteria from NO stress. Nitric oxide (NO) is a lipophilic, freely diffusible radical that can inhibit enzymes, damage DNA, initiate lipid peroxidation, and exacerbate peroxide-induced damage (49,53,66). NO chemistry can be divided into those reactions that occur between NO and biomolecules (direct effects) and those reactions that can only occur subsequent to NO reacting with oxygen or superoxide to form reactive nitrogen oxide species (RNOS) (indirect effects). Direct effects of NO include the formation of metal-nitrosyl complexes (63) and reactions with lipid-derived (50) and other high-energy radicals (34). For example, NO coordinates free or enzyme-bound Fe(II) to form Fe-NO as described for cytochrome P450 (36, 43, 64). Fe nitrosylation leads to altered activity of at least two bacterial metalloregulatory proteins: Fur and Fnr (13,14). Indirect effects occur after NO reacts with either oxygen to generate N 2 O 3 or superoxide to generate the highly reactive oxidant peroxynitrite (OONO Ϫ ). Peroxynitrite rapidly decomposes to form nitrate (NO 3 Ϫ ), hydroxyl radical ( ⅐ OH), and nitrogen dioxide radical ( ⅐ NO 2 ). Some RNOS have the propensity to react with thiol groups and amines to form S-nitrosothiols and nitrosamines.Bacteria encounter NO from a variety of sources. Macrophages of the mammalian immune system generate NO with an inducible NO synthase as part of their arsenal employed against microbial pathogens (38,57). NO synthases have also been identified in some bacteria, including Bacillus subtilis (1, 2), although their function in many cases remains elusive.Denitrifying bacteria produce NO as an intermediate in the reduction pathway from nitrate to dinitrogen (33). B. subtilis is not capable of denitrification, yet it coexists with denitrifying bacteria in subsurface environments. It is therefore likely that B. subtilis has developed a targeted response to exogenously and perhaps endogenously produced NO.Several bacterial enzymes can alleviate NO stress. The Escherichia coli flavohemoglobin Hmp has NO reductase activity under anaerobic conditions (31) and NO dioxygenase (22) or denitrosylase activity (25) under aerobic conditions; however, only the aerobic Hmp activities appear to confer NO stress resistance (20). In addition, both E. coli and Salmonella enterica hmp mutants are hypersensitive to NO (41,56,57). Nakano showed that B. subtilis hmp is regulated by ResDE, a two-comp...

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SummaryWe have used DNA microarrays to monitor the global transcriptional response of Bacillus subtilis to changes in manganese availability. Mn(II) leads to the MntR-dependent repression of both the mntH and mntABCD operons encoding Mn(II) uptake systems.