ARGOS (“Acoustic Reporter Gene Overhaul in Saccharomyces”)

As acoustic reporter genes (ARGs), gas vesicles (GVs) have recently seen a rapid rise in biomedical interest with applications including non-invasive diagnostics and treatment. Gas filled protein nanostructures, GVs are unique, genetically encodable microbubbles that provide a stable ultrasound contrast handle. Due to their complex assembly inside of cells, the use of GVs is currently constricted to a number of wild-type, minimally engineered gene clusters and the range of cells they can be deployed to is limited. By contrast, fluorescent proteins (FPs) as optical reporter genes can readily be deployed to any biological context and decades of protein engineering have imbued them with an expansive array of colors, improved brightness, and sophisticated applications.In this action, we aim to parallel the immense development achieved for FPs and overcome the limitations of re-engineering gas vesicles by linking their successful assembly in yeast cells to a clear optical readout, unlocking high throughput screening by FACS. Utilizing this platform, we will screen libraries of modularly assembled GV gene clusters for successful expression in S. cerevisiae. The Sc2.0 synthetic yeast genome platform will then enable the evolution of yeast genomes for superior GV assembly pathways using SCRaMbLE. Our novel GV gene clusters will be targeted for carefully guided directed (co-)evolution of one or multiple GV genes to develop synthetic GVs that provide improved contrast and phenotypes to expand their applications. To facilitate the swift engineering of ARGs for novel applications, we will obtain next-generation sequencing and single-cell RNA sequencing data of our GV genes and evolved yeast genomes to develop a deeper sequence to function understanding of GV expression in eukaryotes. This action, ARGOS, “Acoustic Reporter Gene Overhaul in Saccharomyces”, will prepare the next generation of GVs for researchers to peer beyond the limits of optical imaging.