HIV-l Vpr blocks host cell cycle progression and interrupts cytoskeletal structures, two effects which may be related to HIV pathogenesis. At present, little is known about how Vpr affects these cellular functions and our studies are designed specifically to address this question. We have developed a novel fission yeast model system, in which we found that Vpr-induced cellular changes were very similar to those observed in human cells. As compared with human systems, fission yeast offers a simple yet genetically well-defined eukaryotic system to study mechanisms of Vpr action at both genetic and biochemical levels. We plan to use wild-type and previously characterized mutants to identify cellular pathways on which Vpr impinges and to discover the function of Vpr, something that cannot yet be done easily in mammalian cells. The fact that p34cdc2, cyclin B and other cell cycle control genes are interchangeable between human and fission yeast makes the fission yeast system relevant for studying related functions. Our goal is to use this model system l) to define domains of Vpr required to interrupt cellular functions; 2) to identify which cell cycle G2 control pathway(s) Vpr affects, and 3) to determine what gene product(s) Vpr interacts with. Our preliminary studies indicated that Vpr is likely interacting with upstream regulators of the G2-M transition but not with gene products from the DNA damage pathway. We have also identified protein candidates that bind to Vpr. In this study, we will pinpoint the cellular targets of the Vpr effects and characterize the Vpr-interacting proteins. We will further expand our studies to test the obtained results in a mammalian system. It is our expectation that the proposed studies will provide new insights into the mechanism of action of Vpr on affecting host cell functions and HIV pathogenesis.
