Head: Mary Lilly, Ph. D.
We use Drosophila oogenesis as a model to explore the developmental regulation of the cell cycle. The long-term goal of the laboratory is to understand how the cell cycle program of the Drosophila ovarian cyst is coordinated with the developmental events of oogenesis. In Drosophila the oocyte develops within the context of a 16-cell germline cyst. Individual cells within the cyst are referred to as cystocytes and are connected by actin-rich ring canals. The cyst is produced through a series of four synchronous mitotic divisions during which cytokinesis is incomplete. While all 16 cystocytes enter premeiotic S phase, only a single cell remains in the meiotic cycle and becomes the oocyte. The other 15 cells enter the endocycle and develop as highly polyploid nurse cells. Currently, we are working to understand how cells within ovarian cyst enter and maintain either the meiotic cycle or the endocycle. In addition, we are examining how this cell cycle choice influences the nurse cell/oocyte fate decision.
Inhibiting Cdk1 activity during prophase of meiosis I.
Animal oocytes inhibit the activity of the mitotic kinase Cdk1 at the onset of meiosis, but must be able to reactivate the kinase much later in oogenesis, as they undergo meiotic maturation and enter the first meiotic division. How cells within early ovarian cysts initiate the long-term inhibition of Cdk1 activity is poorly understood. As is true in many animal oocytes, Drosophila oocytes inhibit the accumulation of the mitotic cyclins, the activating subunits of Cyclin-dependent kinase 1 (Cdk1), via a posttranscriptional mechanism. We have found that the RNA binding protein Bruno inhibits the translation of the mitotic cyclin, cyclin A, during prophase of meiosis I.In the absence of Bruno, ovarian cysts enter meiosis but rapidly accumulate high levels of Cyclin A protein and return to the mitotic cycle. Thus, Bruno is required for the maintenance of meiotic quiescence during prophase of meiosis I. Currently, we are performing genetic screens to identify additional upstream activators and downstream effectors of Bruno mediated Cyclin A translational repression during meiosis. Moreover, we are working to identify additional pathways that regulate Cdk1 activity during early oogenesis.
Preventing DNA replication in prophase I oocytes.
To protect genome integrity the prophase I oocyte must guard against inappropriate DNA replication. Indeed, if the oocyte fires even a single DNA replication origin it is unlikely to produce a functional gamete. In Drosophila, the oocyte is further challenged by its physical connections to nurse cells undergoing repeated rounds of DNA replication. We have provided an explanation for how oocytes prevent inappropriate DNA replication during meiosis I. Specifically, we determined that high levels of the Cyclin-Dependent kinase inhibitor (CKI) Dacapo inhibit DNA replication in the oocyte by preventing the activation of the S phase kinase, Cyclin-dependent kinase 2 (Cdk2). Additionally, our results indicate that restraining Cyclin E/Cdk2 activity is essential for many of the downstream events of oocyte differentiation. Intriguingly, recent evidence suggests that the Dacapo homolog, p27Kip1, may play an important role in regulating growth and cell cycle progression in prophase I arrested mouse oocytes.
Establishing the nurse cell endocycle within ovarian cysts.
As part of exploration of cell cycle regulation and cellular differentiation within ovarian cysts, we are working to delineate the nature of the cell cycle inputs required to run the nurse cell endocycle. These studies have resulted in two major findings. First we have shown that the Anaphase Promoting Complex (APC)/CFzr/cdh1 is essential for endocycle progression. Specifically, we find that APC/CFzr/Cdh1 is required to reduce the levels of the mitotic cyclins and Geminin in order to facilitate the relicensing of DNA replication origins, and cell cycle progression, during the endocycle. Additionally, we have determined that the oscillations of the CKI Dacapo promote endocycle progression by reinforcing low Cyclin E/Cdk2 activity during the endocycle Gap phase. Together this work has helped define the nature of the cell cycle oscillator that drives endocycle progression in Drosophila.