Zinc Binding Inhibits Cellular Uptake and Antifungal Activity of Histatin-5 in Candida albicans

Abstract

) is a polycationic, histidine-rich antimicrobial peptide with potent antifungal activity against the opportunistic fungal pathogen Candida albicans. Hist-5 can bind metals in vitro, and metals have been shown to alter the fungicidal activity of the peptide. Previous reports on the effect of Zn 2+ on Hist-5 activity have been varied and seemingly contradictory. Here, we present data elucidating the dynamic role Zn 2+ plays as an inhibitory switch to regulate Hist-5 fungicidal activity. A novel fluorescently labeled Hist-5 peptide (Hist-5*) was developed to visualize changes in internalization and localization of the peptide as a function of metal availability in the growth medium. Hist-5* was verified for use as a model peptide and retained antifungal activity and mode of action similar to native Hist-5. Cellular growth assays showed that Zn 2+ had a concentration-dependent inhibitory effect on Hist-5 antifungal activity. Imaging by confocal microscopy revealed that equimolar concentrations of Zn 2+ kept the peptide localized along the cell periphery rather than internalizing, thus preventing cytotoxicity and membrane disruption. However, the Zn-induced decrease in Hist-5 activity and uptake was rescued by decreasing the Zn 2+ availability upon addition of a metal chelator EDTA or S100A12, a Zn-binding protein involved in the innate immune response. These results lead us to suggest a model wherein commensal C. albicans may exist in harmony with Hist-5 at concentrations of Zn 2+ that inhibit peptide internalization and antifungal activity. Activation of host immune processes that initiate Zn-sequestering mechanisms of nutritional immunity could trigger Hist-5 internalization and cell killing.

The Synergy Between Zinc and Antimicrobial Peptides: An Insight into Unique Bioinorganic Interactions

“…Though, a similar study by these authors did observe a rescuing effects by Hst5 for a streptococcal strain with impaired Zn(II) efflux within a buffer that mimicked saliva [71]. Reviewing this literature as a whole suggests that Hst5 may have a greater role in developing the microbial composition within an oral cavity by exerting selective antimicrobial activity and maintaining commensal forms of the microbiota, possibly being dictated through signaling of Zn(II) concentrations [44,71,72].…”

“…However, several recent lines of evidence suggest that Zn(II) mitigates the bioactivity of Hst5 in a stoichiometric-dependent manner. At low Zn(II):Hst5 ratios of 0.5:1 or less, Zn(II) was shown to enhance the antifungal activity of Hst5 against C. albicans [44,64]. In fact, fungal cells that were treated with Hst5 and sub-stoichiometric concentrations of Zn(II) were shown to be less virulent when compared with untreated cells, a result of Zn(II)-induced reorganization of the fungal cell wall and impaired adhesion to host epithelial tissues [64].…”

“…Hst5 is generally thought to enter the cytoplasm in an energydependent manner of target microbes, for example via non-specific, membrane-bound polyamine transporters, Dur3 and Dur31, in C. albicans. Once inside the cytoplasm, Hst5 it is thought to cause cell death through generation of reactive oxygen species, alterations in mitochondrial functions, and non-lytic potassium and ATP efflux [44,61,64].…”

Salivary Antimicrobial Peptide Histatin-5 Does Not Display Zn(II)-Dependent or -Independent Activity against Streptococci

“…However, a recent report indicates that the role of Zn­(II) is concentration-dependent: low concentrations of added Zn­(II) enhance the antimicrobial activity of Hst5 against C. albicans (compared with the control without any added Zn­(II)), while high concentrations of added Zn­(II) suppress it …”

“…However, a recent report indicates that the role of Zn(II) is concentration-dependent: low concentrations of added Zn(II) enhance the antimicrobial activity of Hst5 against C. albicans (compared with the control without any added Zn(II)), while high concentrations of added Zn(II) suppress it. 22 Beyond histatins and Zn(II)-binding metallo-peptides, Zn(II)-dependent host innate immune responses are well described. In response to microbial infection, Zn(II) levels and those of Zn(II)-binding or Zn(II)-transporting proteins within a host organism can rise and fall, leading to fluctuations in Zn(II) availability within different niches in the infected host.…”