Characters of homogentisate oxygenase gene mutation and high clonality of the natural pigment-producing Vibrio choleraestrains

Abstract

BackgroundSome microorganisms can produce pigments such as melanin, which has been associated with virulence in the host and with a survival advantage in the environment. In Vibrio cholerae, studies have shown that pigment-producing mutants are more virulent than the parental strain in terms of increased UV resistance, production of major virulence factors, and colonization. To date, almost all of the pigmented V. cholerae strains investigated have been induced by chemicals, culture stress, or transposon mutagenesis. However, during our cholera surveillance, some nontoxigenic serogroup O139 strains and one toxigenic O1 strain, which can produce pigment steadily under the commonly used experimental growth conditions, were obtained in different years and from different areas. The genes VC1344 to VC1347, which correspond to the El Tor strain N16961 genome and which comprise an operon in the tyrosine catabolic pathway, have been confirmed to be associated with a pigmented phenotype. In the present study, we investigated the mechanism of pigment production in these strains.ResultsSequencing of the VC1344, VC1345, VC1346, and VC1347 genes in these pigmented strains suggested that a deletion mutation in the homogentisate oxygenase gene (VC1345) may be associated with the pigmented phenotype, and gene complementation confirmed the role of this gene in pigment production. An identical 15-bp deletion was found in the VC1345 gene of all six O139 pigment-producing strains examined, and a 10-bp deletion was found in the VC1345 gene of the O1 strain. Strict sequence conservation in the VC1344 gene but higher variance in the other three genes of this operon were observed, indicating the different stress response functions of these genes in environmental adaption and selection. On the basis of pulsed-field gel electrophoresis typing, the pigment-producing O139 strains showed high clonality, even though they were isolated in different years and from different regions. Additionally all these O139 strains belong to the rb4 ribotype, which contains the O139 strains isolated from diarrheal patients, although these strains are cholera toxin negative.ConclusionDysfunction of homogentisate oxygenase (VC1345) causes homogentisate accumulation and pigment formation in naturally pigmented strains of V. cholerae. The high clonality of these strains may correlate to an environmental survival advantage in the V. cholerae community due to their pigment production, and may imply a potential protective function of melanin in environmental survival of such strains.

Characterization of the RnfB and RnfG Subunits of the Rnf Complex from the Archaeon Methanosarcina acetivorans

“…The results suggest electron transport from cytochrome c to MP that donates electrons to HdrDE. It is postulated that MP is a potential coupling site for the translocation of protons analogous to the role of MP in the pathway of methanol conversion to methane [2], [9]. The combined evidence supports a H 2 -independent membrane-bound electron transport chain originating with Fd and culminating with reduction of CoM-S-S-CoB involving the Rnf complex, cytochrome c and MP.…”

“…An electron transport chain is proposed for M. acetivorans , the only non-H 2 -metabolizing Methanosarcina species for which the genome is sequenced [9]. The genome does not encode Ech hydrogenase [10], further excluding H 2 in electron transport.…”

“…The cytochrome c is synthesized at high levels in acetate-grown cells where it dominates the UV-visible spectrum of purified membranes [11]. Reduction of purified membranes with Fd from acetate-grown cells leads to reduction of the cytochrome c that is re-oxidized by the addition of either CoM-S-S-CoB or 2-hydoxyphenazine, an analog of MP [9]. Reduced 2-hydoxyphenazine is re-oxidized by membranes dependent on the addition of CoM-S-S-CoB.…”

“…Clearly, organisms with diverse metabolisms are expected to have evolved specialized biochemical properties of Rnf complexes. The complex from M. acetivorans stands apart from characterized Rnf complexes based on its function in methanogenesis, inability to reduce NADH or NADPH [9], [14], reduction of the electron acceptor MP and association with cytochrome c [9], [11]. Previous proposals for the roles of subunits have relied exclusively on bioinformatic analyses and extrapolation from characterized NAD-dependent Rnf complexes of non-methanogenic species classified in the domain Bacteria [2], [11].…”

A Multienzyme Complex Channels Substrates and Electrons through Acetyl-CoA and Methane Biosynthesis Pathways in Methanosarcina

“…Methanogens which solely utilize the hydrogenotrophic pathway have electron transport systems that are different from the electron transport systems of generalist methanogen species like Methanosarcina acetivorans . The generalist organism M. acetivorans is capable of using the methylotrophic (methanol, methylamines, methylsulfides), carboxidotrophic (CO), and the acetoclastic pathways, but cannot use the hydrogenotrophic or methyl respiration pathways due to the lack of expression of suitable hydrogenases [5], [6], [7], [8].…”