This proposal will determine the molecular basis for adhesion of fungi, Saccharomyces and Candida species to inert surfaces. Our goal is to solve both basic and clinically relevant problems as fungal pathogens gain entry into the body by adhesion to indwelling devices. Prevention of adhesion to these devices could stem the increasing prevalence of fungal disease. Adhesion in fungi is conferred by a family of genes that encode cell surface proteins. The basic studies take advantage of our ability to compare the functions of all the genes in the genomes of two closely related strains of S. cerevisiae (S288c and sigma) with a specific emphasis on those genes that affect adhesion. This comparison is possible because the genome of sigma will be sequenced and annotated in concert with the transfer of all the deletion set (~6000 knockouts) from the already sequenced S288c into sigma. Reciprocally, the library of S288c deletions has been transformed with FLO8+ to permit assessment of adhesion in this background. This comparison will reveal all genes required for the adhesion phenotype, and also provide evidence for the regulatory divergence between individuals within an interbreeding population. Variegation, the on/off switching of telomeric and non-telomeric adhesin genes, will be studied by continuous fluorescence microscopy. The cis-acting sequences and trans-acting complexes that control expression and variegation will be analyzed. This analysis will reveal the components of required for establishment and maintenance of this epigenetic state. The control of gene expression by antisense RNA and small open reading frames in the promoters of genes will be determined using bioinformatics as well as molecular genetics. This information obtained from Saccharomyces is used to create strains of C. albicans for use in devising dextran gels containing non-diffusible amphotericin. The Amphogels are effective in an animal model. New work will devise gels that are effective against adherent, non-adherent, and filamentous forms of C. albicans and other fungal species.
Public Health Relevance Statement Fungi, a major source of human disease, gain entry into the body mainly by adhering to medical devices such as catheters. The research proposed will determine the mechanism of adherence by fungi. This basic information will guide the creation of antibiotic containing materials that can be used to coat devices so that the fungi are killed upon contact before they can hitch a ride into the body.
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