Localization of Phr1p in Candida albicans

PHR1 is a pH-regulated gene belonging to a family of five genes of Candida albicans. PHR1 expression is triggered at pH values of the growth medium > 5.5. Loss of PHR1 induces morphological defects, inability to support hyphal growth and reduced virulence in animal mouse models of infection. Phr1p is endowed of a beta(1,3)-glucan elongase activity similarly to Gas1p of S.cerevisiae and Gel1-Gel2 proteins of A. fumigatus. In order to explore the function of Phr1p in Candida albicans morphogenesis we obtained a GFP fusion to determine its cellular localization. During growth in yeast form Phr1p-GFP was predominantly detected at discrete sites of the plasma membrane. Moreover Phr1p-GFP was recovered in the Detergent Resistant Membranes indicating its association with the lipid rafts as already described for wild type Phr1p. Next we examined the localization of Phr1p during hyphal growth. Phr1p-GFP was detected at 30 min after a shift of stationary phase cells to condition of induction of hyphal growth. At this time point the fluorescence of Phr1-GFP was very bright and restricted at the tip of the germ tubes. Phr1p remained highly polarized at the apex of the hyphae as the hyphae grew. The protein also gradually distributed along the lateral cell wall as the hyphae elongated. A study of colocalization with chitin indicated that Phr1p is more abundant at the apex where chitin is less abundant. The results suggest that Phr1p is required at the site of maximal growth probably for the incorporation of new beta(1,3)-glucan in the expanding cell wall but is also retained along the hyphae where the cell wall becomes more cross-linked and resistant. Phr1p also showed an additional localization: it was detected along the septum region in vegetative growing cells and as a thin line in the septa of the hyphae indicating a potential role of the protein also at these sites. Acknowledgement: this work was supported by RTN project Cantrain N. 51248 to LP.