LSF AND MAMMALIAN CELL CYCLE PROGRESSION

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The growth of mammalian cells is driven by signal transduction pathways, whose targets include transcription factors that regulate expression of genes critical to cell cycle progression. At one pivotal point in late G1 phase, the restriction point, cellular signals are integrated to determine whether to replicate cellular DNA (progress to S phase) or revert to a resting state. The signalling molecules required for transit from G1 to S include both D and E cyclins complexed with cyclin-dependent kinases (CDKs). Cyclin D/CDKs phosphorylate Rb and activate members of the E2F family of transcription factors, resulting in activation of genes important for cellular DNA synthesis. Little is known, however, about the regulation of other transcription factors that activate additional components of the DNA synthesis machinery, including potential transcription factors that are targeted by cyclin E/CDK at this key point in the cell cycle. Our recent studies have shown that the mammalian transcription factor LSF plays a key role in cell cycle progression by regulating thymidylate synthase (TS) expression at the G1/S transition. Because TS is essential for cell growth, it has been a cancer chemotherapeutic target for over two decades. Likewise, as an essential regulator of TS expression, LSF may ultimately prove a valuable target for cancer therapies. The DNA-binding activity of LSF is enhanced in late G1, via the modification of pre-existing pools of nuclear LSF. In vitro, LSF is an effective substrate for cyclin E/cdk2. An understanding of the signaling pathway that regulates LSF, and the molecular mechanism by which LSF regulates TS expression may therefore illuminate novel signaling events in cell cycle progression in mammalian cells. Specifically, this proposal will address the following: 1. Is LSF phosphorylated by cyclin E/cdk2 during the late G1 phase? 2. Does phosphorylation of LSF by cyclin E/cdk2 (or other relevant modifications) alter the DNA-binding and transcriptional activities of LSF? 3. Is LSF a major target of cdk2 (or another pathway) in progression of cells into S? 4. Does LSF critically regulate genes other than TS during cell cycle progression?