Oxygen-controlled regulation of the flavohemoglobin gene in Bacillus subtilis

Abstract

A gene, hmp, which encodes a ubiquitous protein homologous to hemoglobin was isolated among genes from Bacillus subtilis that are induced under anaerobic conditions. The hmp protein belongs to the family of two-domain flavohemoproteins, homologs of which have been isolated from various organisms such as Escherichia coli, Alcaligenes eutrophus, and Saccharomyces cerevisiae. These proteins consist of an amino-terminal hemoglobin domain and a carboxy-terminal redox active site domain with potential binding sites for NAD(P)H and flavin adenine dinucleotide. The expression of hmp is strongly induced upon oxygen limitation, and the induction is dependent on a two-component regulatory pair, ResD and ResE, an anaerobic regulator, FNR, and respiratory nitrate reductase, NarGHJI. The requirement of FNR and NarGHJI for hmp expression is completely bypassed by the addition of nitrite in the culture medium, indicating that fnr is required for transcriptional activation of narGHJI, which produces nitrite, leading to induction of hmp expression. In contrast, induction of hmp was still dependent on resDE in the presence of nitrite. A defect in hmp in B. subtilis has no significant effect on anaerobic growth.The gram-positive bacterium Bacillus subtilis was shown to grow anaerobically in the presence of nitrate but not nitrite as a terminal electron acceptor (9,11,16,32,34,36,38). Genes involved in nitrate respiration in B. subtilis have been identified, and components of a regulatory pathway governing anaerobic gene expression are proposed in the accompanying paper (27). According to this model, the two-component signal transduction proteins, a histidine kinase, ResE; and a response regulator, ResD (17,38); are required for nitrate respiration in part to activate transcription of fnr, an anaerobic gene regulator, from an fnr-specific promoter upon oxygen limitation (27). FNR thus produced activates the narK-fnr operon from a promoter located upstream of narK, encoding a protein with nitrite extrusion activity (9,27). fnr is also required for the transcriptional activation of the narGHJI operon (9, 16), which encodes proteins homologous to the three subunits of Escherichia coli respiratory nitrate reductase and a polypeptide required for the assembly of the reductase complex (2-4). Putative FNR-binding sites were identified in the promoter region of narK and narGHJI (9). Mutations in narGHJI (16) as well as moaA which functions in the biosynthesis of molybdopterin, a precursor of the molybdenum cofactor of assimilatory and respiratory nitrate reductases (11), abolished nitrate respiration. In an attempt to uncover the regulation of gene expression responding to oxygen limitation, we have screened promoter activities specifically induced under anaerobic conditions. In this paper, we describe the isolation and characterization of one of these promoters. The activity identified is associated with a gene encoding a putative flavohemoglobin-like protein, the expression of which is induced by nitrite and requires the two-component reg...

Vitreoscilla Hemoglobin Binds to Subunit I of Cytochrome bo Ubiquinol Oxidases

“…In any case, it now appears that the roles and functions of VHb are more complex than just the oxygen-transporting one initially believed. These hypothetical functions are different from the NO detoxification function proposed for flavohemoproteins from E. coli (22)(23)(24)(25)(26)(27). A recent study, however, indicates that NO detoxification may be a secondary but important function for VHb (28), whereas its primary function may be oxygen transfer, as supported by the data presented here.…”

Characterization of a New Hemoprotein in the YeastSaccharomyces cerevisiae

“…Although the function of these proteins is still unclear, oxygen appears to play a role in the expression of at least some of them. For example, growth in limiting concentrations of oxygen induces the synthesis of the Vitreoscilla Hb (7) and of the Bacillus subtilis (8) and Alcaligenes eutrophus (9) flavohemoglobins. In contrast, transcription of the Saccharomyces cerevisiae flavohemoglobin, encoded by the YHB1 gene, is enhanced under oxygen-replete conditions via the hemedependent activation of the HAP1 and HAP2/3/4 transcription factors (10).…”

Paraquat regulation of hmp (flavohemoglobin) gene expression in Escherichia coli K-12 is SoxRS independent but modulated by sigma S

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