Viruses infect cells and use the host cell machinery to create more viruses. Some viruses do this by hijacking cellular processes (e.g. cell division cycle) to force cells into states where they grow more and produce more virus. Recent work from the investigators' labs showed that a viral gene (SBF) hijacked and rewired the cell cycle of an ancestor of Fungi and replaced the ancestral regulator (E2F) without disrupting the function of the cell cycle regulatory network. Some Fungi (e.g. Chytrids) have a hybrid network with both ancestral and virally-derived cell cycle genes, which reflects a transition state during the rewiring of the cell cycle. Unfortunately, nothing is known about the Chytrid cell cycle. This project will study the function and architecture of the hybrid cell cycle network in a Chytrid (Spizellomyces punctatus) using genomics. The proposed work is significant because it will provide new data to elucidate how a virus could be a driver of evolution by contributing new genes and regulatory elements into an essential and highly conserved cell cycle network. This project also integrates graduate and post-doctoral research with high school teaching to develop the next generation of scientists.
Part 2: technical
The hypothesis of this proposal is that the viral SBF initially gained cell cycle control by binding cis-regulatory sequences of E2F-responsive promoters, and thereby gained the ability to control the G1/S transition. Early SBF then became redundantly integrated into the ancestral G1/S regulatory network, allowing the dikaryotic ancestor of Fungi to lose the E2F transcription factor. The project will generate data to address the central hypothesis and its alternatives with two specific aims: (1) Characterize the evolution of the DNA-binding specificity of recombinant SBF and E2F in early-diverging Fungi using high-throughput in vitro assays; and (2) Measure gene expression, chromatin accessibility, and transcription factor binding to promoters using genomic methods in cell cycle synchronized cultures of Spizellomyces punctatus. The in vitro data will identify overlapping and/or distinct DNA binding specificities and their conservation across early-diverging Fungi. The Spizellomyces data will identify cell cycle genes and elucidate the cis-regulatory architectures of SBF and E2F-regulated genes in an exemplary Chytrid that has both factors. Conservation of overlapping DNA-binding specificity of SBF and E2F across most early-diverging Fungi and redundant regulation of cell cycle genes in Spizellomyces would support the central hypotheses.
