Killing of Candida albicans by Human Salivary Histatin 5 Is Modulated, but Not Determined, by the Potassium Channel TOK1

Abstract

ABSTRACT Salivary histatin 5 (Hst 5), a potent toxin for the human fungal pathogen Candida albicans , induces noncytolytic efflux of cellular ATP, potassium, and magnesium in the absence of cytolysis, implicating these ion movements in the toxin's fungicidal activity. Hst 5 action on Candida resembles, in many respects, the action of the K1 killer toxin on Saccharomyces cerevisiae , and in that system the yeast plasma membrane potassium channel, Tok1p, has recently been reported to be a primary target of toxin action. The question of whether the Candida homologue of Saccharomyces Tok1p might be a primary target of Hst 5 action has now been investigated by disruption of the C. albicans TOK1 gene. The resultant strains ( TOK1/tok1 ) and ( tok1/tok1 ) were compared with wild-type Candida ( TOK1/TOK1 ) for relative ATP leakage and killing in response to Hst 5. Patch-clamp measurements on Candida protoplasts were used to verify the functional deletion of Tok1p and to provide its first description in Candida . Tok1p is an outwardly rectifying, noisily gated, 40-pS channel, very similar to that described in Saccharomyces . Knockout of CaTOK1 ( tok1/tok1 ) completely abolishes the currents and gating events characteristic of Tok1p. Also, knockout ( tok1/tok1 ) increases residual viability of Candida after Hst 5 treatment to 27%, from 7% in the wild type, while the single allele deletion ( TOK1/tok1 ) increases viability to 18%. Comparable results were obtained for Hst-induced ATP efflux, but quantitative features of ATP loss suggest that wild-type TOK1 genes function cooperatively. Overall, very substantial killing and ATP efflux are produced by Hst 5 treatment after complete knockout of wild-type TOK1 , making clear that Tok1p channels are not the primary site of Hst 5 action, even though they do play a modulating role.