At the present time, our work focuses on cell cycle regulation by a mammalian form of C. elegans lin-9. In C. elegans, lin-9 functions downstream of the mammalian equivalent of CDK4 in a pathway that regulates cell proliferation. We demonstrate that this lin-9 ortholog, referred throughout the proposal as BARA/LIN-9, also appears to act downstream of cyclin D/CDK4 in mammalian cells. This is supported by the fact that over- expression of BARA/LIN-9 has an inhibitory effect on cell proliferation which is partially blocked by co- expression of cyclin D1 and, furthermore, the partial rescue of phenotypic alterations in CDK-null mice by expression of a mutant form of BARA/LIN-9 lacking amino acids 1-84 (BARA/LIN-9?84). Recently, we have determined that BARA/LIN-9 interacts with p107/p130, members of the pocket protein family, and E2F4. These preliminary results are novel and extremely important in further illuminating our understanding of cell cycle regulation. Obviously, it is through alterations, typically through genetic modifications, of normal cell cycle function that leads to an unchecked state of cell growth, which ultimately promotes tumor formation. It is my intention to pursue this study further as I begin my transition from mentored postdoctoral fellow to independent investigator. To that end, this proposal, if funded, would allow me to pursue a more detailed study of the protein interactions between BARA/LIN-9, p107/p130 and E2F4 through studies that will involve protein mapping with recombinant proteins. Furthermore, I will study the stability of this complex temporally along the cell cycle and examine the potential interaction with other cell cycle proteins. Changes of this complex along different phases of the cell cycle will help us to better understand the potential signaling mechanisms that may regulate this protein complex. In addition, I will also use primary mouse fibroblasts (MEFs) from mice expressing BARA/LIN-9?84 to determine if the interactions with p107/p130 and E2F4 are altered. Moreover, I will continue to study the knockout (KO) mice we have generated to elucidate a mechanism for cyclin D/CDK4 regulation of LIN-9 function. Taken together, this study will produce a more enhanced understanding of cell cycle regulation, which will add substantially to our continuously growing knowledge base and aid the entire cancer research field as we continuously look for novel signaling mechanisms that may translate into new therapeutic approaches in the clinical field. [unreadable] [unreadable] [unreadable]