Mitochondrially driven viral innate immunity in budding yeast

点击翻译按钮获取中文摘要

S. cerevisiae is chronically infected with RNA viruses that rely on mitotic transmission for their persistence. We have discovered that the yeast homolog of mitochondrially localized endonuclease G, Nuc1, opposes the deleterious accumulation of these viruses in meiotic cells through an apparent ancestral cell death pathway. The most widely studied yeast viruses are the double stranded (ds) RNA virus L-A and its frequent “Killer” satellite viruses. The L-A genome encodes gag and gag-pol fusion proteins that assemble into virus-like-particles that support L-A propagation. Varied dsRNA “Killer” viruses exist as satellites of L-A. The most common is “M”, which encodes a processed and secreted toxin called K1 that kills neighboring cells lacking immunity, which is conferred by the Killer genome itself. This proposal will describe investigations of an innate immune function for Nuc1 as well as of the mysterious relationship between L-A and the yeast host. Yeast gametogenesis culminates in the development of 4 meiotic products within the remnant of the cell that produced them. We showed that programmed cell death (PCD) of this remnant occurs as an intrinsic aspect of sporulation and is executed through vacuolar rupture. Intriguingly, NUC1 is required for the fragmentation of genomic DNA from meiotic products frequently swept up in this PCD. This provocative association has remained puzzling however, as PCD proceeds unperturbed in the absence of NUC1. Notably, this is also the case with metazoan apoptosis and its associated DNA fragmentation factors (including endonuclease G), which have no known instructive role in PCD. Indeed, despite the ubiquitous association of mitochondrially driven DNA fragmentation pathways with diverse forms of PCD in multiple kingdoms of life, nothing is known of any adaptive roles they may play. We have now determined that NUC1 functions in a rudimentary apoptotic module that protects sporulating yeast from lethal viral accumulation. Via a meiotically programmed transition, Nuc1 is released from mitochondria and accumulates in the cytosol in a manner dependent on the Por1/2 voltage dependent anion channel (VDAC), which is known to similarly mediate release of mitochondrial content during apoptosis. In the cytosol, Nuc1 functions to repress the accumulation of L-A and Killer dsRNA viruses. This developmentally programmed apoptotic-like process collaborates with the well-established yeast “innate immunity” SKI complex (“Super-Killer”) to prevent a lethal accumulation of the Killer virus encoded toxin in the meiotic progeny. This proposal will advance these findings in three ways. We will examine the NUC1 anti-viral mechanism(s), address the regulation and nature of Por1/2 mediated Nuc1 release, and identify additional targets of Nuc1 during gametogenesis. These will involve continued hypothesis driven genetic, cell biological, and biochemical experiments as well as a simple but powerful genomic screening strategy.