Functional analysis of Candida albicans genes whose Saccharomyces cerevisiae homologues are involved in endocytosis

Abstract

PCR-based techniques for directed gene alterations have become standard tools in Candida albicans. To help to increase the speed of functional analysis of Candida albicans genes, we previously constructed and updated a modular set of pFA-plasmid vectors for PCR-based gene targeting in C. albicans. Here we report the functional analyses of C. albicans ORFs whose homologues in S. cerevisiae are involved in endocytosis, to explore their potential involvement in polarized cell growth. Three C. albicans genes, ABP1, BZZ1 and EDE1, were found to be non-essential. Yeast and hyphal morphogenesis were not affected by the individual deletions and the mutant strains appeared wild-type-like under the different growth conditions tested. On the other hand, deletion of both alleles of the C. albicans PAN1 homologue was not feasible. Promoter shut-down experiments using a MET3p-PAN1/pan1 strain indicated severe growth defects and abolished endocytosis, indicating that PAN1 is an essential gene. Subcellular distribution of CaAbp1 and CaPan1 was analysed via GFP-tagged proteins. Both proteins were found to localize at the cortex and at hyphal tips in a patch-like manner, supporting their role in endocytosis. Localization patterns of Abp1 and Pan1, however, were distinct from that of the FM4-64 stained Spitzenkörper.