Affiliation:
1. Department of Microbial and Molecular Ecology, Institute of Life Sciences, The Hebrew University of Jerusalem, 91904 Jerusalem, Israel
2. Institut National de la Santé et de la Recherche Médicale (INSERM U411), CHU Necker Enfants-Malades, 75730 Paris Cedex 15, France
Abstract
ABSTRACT
Vibrio cholerae
, the causative agent of cholera, is a normal inhabitant of aquatic environments, where it survives in a wide range of conditions of pH and salinity. In this work, we investigated the role of three Na
+
/H
+
antiporters on the survival of
V. cholerae
in a saline environment. We have previously cloned the Vc-
nhaA
gene encoding the
V. cholerae
homolog of
Escherichia coli
. Here we identified two additional antiporter genes, designated Vc-
nhaB
and Vc
-nhaD
, encoding two putative proteins of 530 and 477 residues, respectively, highly homologous to the respective antiporters of
Vibrio
species and
E. coli.
We showed that both Vc-NhaA and Vc-NhaB confer Na
+
resistance and that Vc-NhaA displays an antiport activity in
E. coli
, which is similar in magnitude, kinetic parameters, and pH regulation to that of
E. coli
NhaA. To determine the roles of the Na
+
/H
+
antiporters in
V. cholerae
, we constructed
nhaA
,
nhaB
, and
nhaD
mutants (single, double, and triple mutants). In contrast to
E. coli
, the inactivation of the three putative antiporter genes (Vc-
nhaABD)
in
V. cholerae
did not alter the bacterial exponential growth in the presence of high Na
+
concentrations and had only a slight effect in the stationary phase. In contrast, a pronounced and similar Li
+
-sensitive phenotype was found with all mutants lacking Vc-
nhaA
during the exponential phase of growth and also with the triple mutant in the stationary phase of growth. By using 2-
n
-nonyl-4-hydroxyquinoline
N
-oxide, a specific inhibitor of the electron-transport-linked Na
+
pump NADH-quinone oxidoreductase (NQR), we determined that in the absence of NQR activity, the Vc-NhaA Na
+
/H
+
antiporter activity becomes essential for the resistance of
V. cholerae
to Na
+
at alkaline pH. Since the ion pump NQR is Na
+
specific, we suggest that its activity masks the Na
+
/H
+
but not the Li
+
/H
+
antiporter activities. Our results indicate that the Na
+
resistance of the human pathogen
V. cholerae
requires a complex molecular system involving multiple antiporters and the NQR pump.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
74 articles.
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