Zinc binding inhibits cellular uptake and antifungal activity of Histatin-5 in Candida albicans

Abstract

Histatin-5 (Hist-5) is a polycationic, histidine-rich antimicrobial peptide with potent antifungal activity against the opportunistic fungal pathogen Candida albicans. Hist-5 has the ability to bind metals in vitro and metals have been shown to alter the fungicidal activity of the peptide. Previous reports on the effect of Zn2+ on Hist-5 activity have been varied and seemingly contradictory. Here we present data elucidating the dynamic role Zn2+ 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 Zn2+ had a concentration-dependent inhibitory effect on Hist-5 antifungal activity. Imaging by confocal microscopy revealed that equimolar concentrations of Zn2+ 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 Zn2+ 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 Zn2+ 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.

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. 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.…”

“…These antagonistic host responses, known as "nutritional immunity", 23 suppress microbial growth in the host and inhibit the progress of infectious disease. Although Zn(II) influences the activity of Hst5, 22 it is unclear whether histatins themselves participate in nutritional immunity by modulating Zn(II) availability to microbes.…”