Presentation description

The ESCRT pathway is necessary for essential cell functions such as cytokinesis and vesicle formation. Evolutionary conservation of ESCRTs across eukaryotes underscores the importance of these core functions. However, enveloped viruses take advantage of this by hijacking the pathway in viral budding. Our lab discovered retroCHMP3 in South American primates and house mice, which selectively inhibits viral budding while permitting host ESCRT functions. RetroCHMP3 is a retrocopy of the ESCRT subunit, CHMP3, with a ~70 amino acid c-terminus truncation and ~22 amino acid substitutions. Interestingly, truncation of CHMP3 alone is cytotoxic - only with these mutations does retroCHMP3 become tolerable to the host. However, the steps for detoxification and how essential these mutations are remains unknown. In this project, we used MEDUSA, an established yeast model for directed evolution of ESCRT proteins, to experimentally evolve truncated CHMP3 and ask the question if it can overcome cytotoxicity. Here, we evolved wild type and VPS24 KO (the yeast CHMP3 ortholog) yeast lines transformed with truncated CHMP3 in MEDUSA. Cultures were sampled daily for sequencing and analysis. By day 17, VPS24 KO constructs showed increased expression of truncated CHMP3, while wild types showed decreased expression. This suggests that the VPS24 KO contructs acquired beneficial adaptations over this time.