Adhesin Amyloid Biology

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Heterotypic interactions between amyloid proteins are critical in understanding the outcomes of amyloid-based infection and treatment; however, the molecular and structural determinants that allow or limit amyloid cross- seeding have been difficult to define. Towards this goal, it will be informative to identify amyloid interactions on a larger scale, generating a knowledgebase that can contribute to the identification of rules governing cross- seeding among amyloids. The adhesins are an important family of functional amyloids central to fungal biology and virulence. Adhesins are cell wall-attached proteins that mediate cell-cell adhesion in fungal filamentous growth and biofilm formation. Notably, nearly 90% of adhesins are predicted functional amyloids, and many have been demonstrated to possess core sequences capable of forming amyloid fibrils in solution. In the yeasts S. cerevisiae and C. albicans, adhesins aggregate to form cell surface nanodomain patches important in the enhanced cell-cell adhesion of pseudohyphal and hyphal filaments. The ability to transition between yeast-like and filamentous growth forms is required for virulence in the opportunistic human fungal pathogen C. albicans, and adhesins have been identified as virulence factors. Adhesin sequences with strong amyloidogenic potential are required for wild-type cell-cell adhesion and filamentation. Fungi preferentially bind to like cells in biofilms and filamentous communities. Since this binding is mediated through adhesins, we hypothesize that specificity in amyloid interactions underlies kin discrimination and cellular self-recognition in yeast. Our preliminary data are consistent with this hypothesis, identifying important sequence variation in amyloidogenic regions of the well-studied adhesin Flo11p between filamentous and non-filamentous strains of S. cerevisiae. We also identify that single amino acid changes in amyloid proteins can strongly affect cross- seeding specificity. Here, we propose to test adhesin protein isoforms of Flo11p from filamentous and non- filamentous strains of S. cerevisiae for cross-seeding. Species barriers to adhesin interactions will be tested using the set of identified adhesins from S. cerevisiae and C. albicans. The adhesins present an informative platform for the investigation of amyloid interaction specificity, and a genome-wide set of yeast functional amyloids will be tested for amyloidogenic potential and heterotypic interactions using the adhesin framework as a novel screening scaffold. Collectively, this work will determine the specificity and barriers that constrain heterotypic adhesin interactions in yeast, while offering broader insight into the rules of amyloid cross-seeding. Using the adhesins as a scaffold for yeast surface display, we present a method amenable to high throughput applications for the identification of new amyloids and the rapid assessment of specific aggregation barriers. Adhesins are fungal-specific virulence determinants relevant as targets for new antifungal treatments.