KtrAB and KtrCD: Two K+Uptake Systems inBacillus subtilisand Their Role in Adaptation to Hypertonicity

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Abstract
Recently, a new type of K+transporter, Ktr, has been identified in the bacteriumVibrio alginolyticus(T. Nakamura, R. Yuda, T. Unemoto, and E. P. Bakker, J. Bacteriol.180:3491-3494, 1998). The Ktr transport system consists of KtrB, an integral membrane subunit, and KtrA, a subunit peripherally bound to the cytoplasmic membrane. The genome sequence ofBacillus subtiliscontains two genes for each of these subunits:yuaA(ktrA) andykqB(ktrC) encode homologues to theV. alginolyticusKtrA protein, andyubG(ktrB) andykrM(ktrD) encode homologues to theV. alginolyticusKtrB protein. We constructed gene disruption mutations in each of the fourB. subtilis ktrgenes and used this isogenic set of mutants for K+uptake experiments. Preliminary K+transport assays revealed that the KtrAB system has a moderate affinity with aKmvalue of approximately 1 mM for K+, while KtrCD has a low affinity with aKmvalue of approximately 10 mM for this ion. A strain defective in both KtrAB and KtrCD exhibited only a residual K+uptake activity, demonstrating that KtrAB and KtrCD systems are the major K+transporters ofB. subtilis. Northern blot analyses revealed thatktrAandktrBare cotranscribed as an operon, whereasktrCandktrD, which occupy different locations on theB. subtilischromosome, are expressed as single transcriptional units. The amount of K+in the environment or the salinity of the growth medium did not influence the amounts of the variousktrtranscripts. A strain with a defect in KtrAB is unable to cope with a sudden osmotic upshock, and it exhibits a growth defect at elevated osmolalities which is particularly pronounced when KtrCD is also defective. In thektrABstrain, the osmotically mediated growth defect was associated with a rapid loss of K+ions from the cells. Under these conditions, the cells stopped synthesizing proteins but the transcription of the osmotically inducedproHJ,opuA, andgsiBgenes was not impaired, demonstrating that a high cytoplasmic K+concentration is not essential for the transcriptional activation of these genes at high osmolarity. Taken together, our data suggest that K+uptake via KtrAB and KtrCD is an important facet in the cellular defense ofB. subtilisagainst both suddenly imposed and prolonged osmotic stress.