LSF AND MAMMALIAN CELL CYCLE PROGRESSION

哺乳动物细胞的生长是由信号转导途径驱动的，其靶点包括调节对细胞周期进程至关重要的基因表达的转录因子。在G1期晚期的一个关键点，即限制点，细胞信号被整合以确定是否复制细胞DNA（进展到S期）或恢复到静息状态。从G1期到S期所需的信号分子包括与细胞周期蛋白依赖性激酶（CDK）复合的D和E细胞周期蛋白。 细胞周期蛋白D/CDK磷酸化Rb并激活转录因子E2F家族的成员，导致对细胞DNA合成重要的基因的激活。 然而，很少有人知道，其他转录因子的调节，激活额外的组件的DNA合成机制，包括潜在的转录因子，细胞周期蛋白E/CDK在细胞周期的这个关键点为目标。我们最近的研究表明，哺乳动物转录因子LSF通过调节G1/S转换期胸苷酸合成酶（TS）的表达在细胞周期进程中起着关键作用。 由于TS是细胞生长所必需的，它已经成为癌症化疗靶点超过二十年。 同样，作为TS表达的重要调节因子，LSF可能最终被证明是癌症治疗的有价值的靶点。 LSF的DNA结合活性在G1晚期通过修饰预先存在的核LSF库而增强。 在体外，LSF是细胞周期蛋白E/cdk2的有效底物。 因此，了解调节LSF的信号通路以及LSF调节TS表达的分子机制可能会阐明哺乳动物细胞细胞周期进展中的新信号事件。具体而言，该提案将解决以下问题：1. LSF在G1期晚期被cyclin E/cdk2磷酸化吗？2.细胞周期蛋白E/cdk2（或其他相关修饰）对LSF的磷酸化是否改变了LSF的DNA结合和转录活性？3. LSF是否是cdk2（或另一种途径）在细胞向S进展中的主要靶点？4. LSF在细胞周期进程中是否对TS以外的基因有重要调控作用？

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?