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Project I
Genetic Analysis of E2F function

The G1-S Transition is a Key Cell Cycle Control Point
Many cells make a decision whether or not to enter into a new cell cycle during G1 phase . Consequently, most growth regulatory inputs are exerted on the cell cycle machinery controlling the G1-S transition. My laboratory uses genetic approaches to explore how the G1-S transition is regulated during Drosophila development, and to determine how key regulators of this transition contribute to cell cycle control. Elucidating these mechanisms will further our knowledge of how growth and cell division are coordinated with morphogenesis during development, and provide mechanistic information useful for understanding mammalian cancer.
In animal cells complexes of D type cyclins with cdk4 or cdk6 and cyclin E/cdk2 regulate entry into S phase. Similarly, in the yeast S. cerevisiae a structurally divergent but analogous set of cyclins (CLNs) bind to and activate a single catalytic subunit, CDC28, to regulate the G1-S transition in response to growth signals. Cyclin/cdk-mediated phosphorylation of key target molecules during G1 is thought to trigger a series of downstream events leading to the initiation of DNA replication and cell cycle progression. Molecules known to respond to cyclin/cdk phosphorylation include specialized transcription factors that regulate the expression of genes required for growth and the execution of DNA synthesis. These transcription factors play an essential role in controlling the G1-S transition in mammals, yeast, and insects. In animal cells, this role is played by a family of transcription factors termed E2F/DP.

E2F/DP and Cell Cycle Control
Numerous transcriptional changes occur during progression through the eukaryotic cell cycle. Many genes are coordinately expressed at only one phase of the cell cycle, and they generally encode factors that are involved in executing the events of that particular cell cycle phase. Consequently, these transcriptional programs are thought to contribute in a major way to progression through the cell cycle. One of the best known examples of such a program occurs at the G1-S transition. Work over the past 15 years in animal systems has pointed to the E2F family of transcription factors as key regulators of entry into S phase. "E2F" is a heterodimeric transcription factor containing a molecule of E2F bound to a molecule of DP. In mammals there are families of E2F and DP proteins that have both unique and redundant functions. E2F/DP heterodimers are regulated mainly by association with a member of the pRB family of tumor suppressor proteins (i.e. pRB, p107 and p130), each of which also performs both unique and redundant functions. In spite of the biological complexity generated by these gene families, a unifying model of how these complexes control the cell cycle has emerged. In the simplest version of the model (Figure 1), E2F is both an activator and repressor of transcription. Hypophosphorylated pRB binds to E2F in quiescent cells, and this complex actively represses genes containing E2F binding sites. These target genes fall into two categories: 1) those that encode the "nuts and bolts" required for DNA synthesis to initiate and proceed (e.g. RNR, pol , MCM, ORC, PCNA), and 2) those that encode direct regulators of cell cycle transitions (e.g. cyclin E, cyclin A, cdk's). When quiescent cells are stimulated to divide by exposure to growth factors, pRB is hyperphosphorylated by newly activated G1 cyclin-cdk kinases and dissociates from E2F. This converts E2F from a transcriptional repressor to a transcriptional activator that induces expression of both classes of target genes, thereby stimulating entry into S phase.
Mouse knockout experiments have provided some evidence for this hypothesis in mammals. However, much of this evidence comes from the analysis of embryonic fibroblasts in culture, and less is known about how the E2F family affects cell cycle progression in various tissues in the intact animal. Drosophila provides a simpler genetic system to examine the contribution of E2F function to cell cycle control directly in a developing animal. The Drosophila genome contains two E2Fs (dE2F and dE2F2), a single dDP, and two pRB (RBF and RBF2) genes. Much of our research program involves a genetic analysis of the Drosophila E2F genes. If you wold like more detail of a couple ongoing projects, click here.