Fungal pathogens such as Candida spp. depend on adhesion proteins to attach to host surfaces and invade tissues. Adhesion is complicated by the 100 nm thick carbohydrate based inner cell wall consisting of chitin and β-glucans, making it necessary for adhesion proteins to reach beyond this barrier. Glycosylphosphatidylinositol (GPI) -anchored proteins undergo modification of their glycolipid tails, and are transferred from the plasma membrane to the cell wall surface, where they are attached by a remnant of the GPI-tail. The pathway of enzymes that facilitates this transfer has been partially identified, and involves some unique families of proteins. We recently described an adhesion assay using physiologic fluid shear in Candida parapsilosis where binding required the Als7 adhesion protein. Here, we present data showing that Phr1, a GH72 family transglycosylase, may play a role in facilitating adhesion. Truncation of Phr1 using CRISPR-HDR leads to loss of adhesion. GH72 family transglycosylases cross link β-1,3-glucans and in cooperation with the GH76 family of α-1,6-mannases may also play a role in cross-linking GPI-anchored cell wall proteins to the cell wall. We have also truncated the two GH76 proteins, Dcw1 and Dfg5, in C. parapsilosis, and are currently testing their role in adhesion using a similar approach. In a second approach, we are generating soluble forms of these proteins for use in in vitro assays of enzyme function. Determination of the pathways responsible for modification of fungal cell wall proteins and their attachment to the carbohydrate cell wall may help identify unique and specific inhibitors of fungal adhesion and pathogenesis. Because of their extracellular location, glycan modifying enzymes in this pathway may offer easy access to potential soluble inhibitors making them attractive targets for development of novel antifungal agents.

Attachment of fungal cell wall proteins to the carbohydrate cell wall

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