Fungal infections are a major cause of morbidity and mortality in immunocompromised patients, and Candida albicans is the major causative fungal agent. Adhesion and colonization are crucial first steps in pathogenesis, and cell wall adhesins of the ALS gene family, including Als1p and AIs5p, mediate both adhesion to extracellular matrix and aggregation of the fungus into mini-colonies. Our long term goal is to understand the mechanisms and role of fungal adhesins in cellular interactions. Our current objective is to delineate the molecular properties and activities of AIs5p. AIs5p binds to diverse peptide ligands and mediates both substrate adhesion and cellular aggregation of Saccharomyces cerevisiae cells with the protein displayed on the surface. Our working hypothesis is that binding of AIs5p to diverse peptide ligands mediates adhesion and triggers a self-propagating conformational shift from a molten-globule-like state to a state characterized by amyloid-like mass association. This conformational shift mediates cell association to form macroscopic aggregates.
Four specific aims, based on preliminary results, will determine whether these two activities can be localized to specific domains of the protein, or alternatively, how the domains interact to mediate adhesion and aggregation.
These aims are tests of specific hypotheses: 1) that adhesion results from binding of the globular N-terminal domain of AIs5p to specific peptide ligands; 2) that the differences in ligand specificities of Als1p and AIs5p are due to differences in sequence of the globular N-terminal regions; 3) that the Thr-rich domains of AIs5p mediate homotypic association, and a test of the effect of the number of repeats; and 4) that adhesion leads to self-propagating global conformational change in AIs5p, resulting in aggregation of cells that express the adhesin. This work will be innovative in that a model of conformational shift to an amyloid-like state has not been applied to cell adhesion. Furthermore, this model involves a widespread type of sequence (Thr-rich) that has not been studied before. The rationale for this work is that it will elucidate a novel cell-adhesion strategy in a common and serious pathogen. The results will be general in that AIs homologs are present in other pathogenic fungi, and similar Thr-rich sequences are found in cell surface proteins in other pathogens including Staphylococcus, Mycobacterium tuberculosis, and Plasmodium.
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