Skip to main content

Home > Section on Gamete Development

Cell Cycle Regulation in Oogenesis

Mary Lilly, PhD
  • Mary Lilly, PhD, Head, Section on Gamete Development
  • Melissa Agnello, BS, Student
  • John Csokmay, MD, Clinical Fellow
  • Eva Decotto, PhD, Visiting Fellow
  • Guillermo Gonzalez, BS, Technician
  • Remmy Kasili, PhD, Visiting Fellow

The long-term goal of the laboratory is to understand how the cell cycle events of meiosis are coordinated with the developmental events of gametogenesis. Chromosome missegregation during female meiosis is the leading cause of miscarriages and birth defects in humans. Recent evidence suggests that many meiotic errors are downstream of defects in oocyte growth and/or the hormonal signaling pathways that drive the differentiation of the oocyte. Thus, an understanding of how meiotic progression and gamete differentiation are coordinated during oogenesis is essential to studies in both reproductive biology and medicine. Currently, we are using the genetically tractable model organism Drosophila melanogaster to examine how meiotic progression is both coordinated with and instructed by the developmental program of the egg.

Regulation of developmentally programmed endocycles

The endocycle is a commonly observed variant cell cycle in which cells undergo repeated rounds of DNA replication with no intervening mitosis. Cells that are highly metabolically active, such as the giant trophoblast of the mammalian placenta and Drosophila ovarian nurse cells, often grow via endoreplication. How the cell cycle machinery is modified to transform a mitotic cycle into endocycle has long been a question of interest. In both plants and animals, the transition from the mitotic cycle to the endocycle requires Fzr/Cdh1, a positive regulator of the anaphase-promoting complex/cyclosome (APC/C). However, because many of its targets are transcriptionally downregulated upon entry into the endocycle, it remained unclear whether the APC/C functioned beyond the mitotic/endocycle boundary. We have shown that APC/CFzr/Cdh1 activity is required to promote the G/S oscillation of the Drosophila endocycle. We find that compromising APC/C activity, after cells have entered the endocycle, inhibits DNA replication and results in the accumulation of multiple APC/C targets including the mitotic Cyclins and Geminin. Notably, our data suggest that the activity of APC/CFzr/Cdh1 during the endocycle is not continuous but cyclic, as demonstrated by the APC/C–dependent oscillation of the pre-replication complex component ORC1. Taken together our data suggest a model in which, during the Drosophila endocycle, the cyclic activity of APC/CFzr/Cdh1 is driven by the periodic inhibition of Fzr/Cdh1 by Cyclin E/Cdk2. Thus, we propose that, as is observed in mitotic cycles, APC/CFzr/Cdh1 functions during endocycles to reduce the levels of the mitotic Cyclins and Geminin in order to facilitate the relicensing of DNA replication origins and cell cycle progression. These studies have allowed us to formulate and test new models on the minimum cell cycle inputs necessary to construct a G/S oscillator.

Meiotic progression and oocyte differentiation in early ovarian cysts

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. 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 the ovarian cyst enter and maintain either the meiotic cycle or the endocycle. In addition, we are examining how this cell cycle choice drives the development of the mature egg.

The proper execution of premeiotic S phase is essential to both the maintenance of genomic integrity and accurate chromosome segregation during the meiotic divisions. However, the regulation of premeiotic S phase remains poorly defined in metazoa. We identified the p21(Cip1)/p27(Kip1)/p57(Kip2)–like cyclin-dependent kinase inhibitor (CKI) Dacapo (Dap) as a key regulator of premeiotic S phase and genomic stability during Drosophila oogenesis. In dap−/− mutant females, ovarian cysts enter the meiotic cycle with high levels of Cyclin E/cyclin-dependent kinase 2 (Cdk2) activity and accumulate DNA damage during the premeiotic S phase. High Cyclin E/Cdk2 activity inhibits the accumulation of the replication-licensing factor Doubleparked/Cdt1 (Dup/Cdt1). Accordingly, we find that dap−/− ovarian cysts have low levels of Dup/Cdt1. Moreover, mutations in dup/cdt1 dominantly enhance the dap−/− DNA-damage phenotype. Importantly, the DNA damage observed in dap−/− ovarian cysts is independent of the DNA double-strand breaks that initiate meiotic recombination. Together, our data suggest that the CKI Dap promotes the licensing of DNA replication origins for the premeiotic S phase by restricting Cdk activity in the early meiotic cycle. We are currently working to define additional regulators of the premeiotic S phase in Drosophila.

The pathways that control entry into the meiotic cycle and early meiotic progression are poorly understood in metazoans. We previously identified a gene, missing oocyte (mio), that regulates nuclear architecture and meiotic progression in early ovarian cysts. In mio mutants, the oocyte enters the meiotic cycle and progresses to pachytene. However, this meiotic state is not maintained, and ultimately the oocyte withdrawals from meiosis, enters the endocycle and becomes polyploid. mio mutants display some of the earliest meiotic defects reported in Drosophila. Moreover, the Mio protein accumulates in the oocyte nucleus in early prophase of meiosis I. Therefore, mio provides an excellent entry point to explore how the unique cell biology of the early ovarian cyst and the establishment of the meiotic program influence the downstream events of oocyte differentiation and meiotic progression. To better understand the role of mio in oogenesis, we initiated a series of experiments to identify additional proteins that function in the Mio pathway. From these studies, we have found that Mio physically and genetically interacts with the nucleoporin Nup44A. Moreover, mutations in nup44A disrupt meiotic progression and cause female sterility. These observations provide the framework for future studies on how nuclear pore components influence meiotic progression and the maintenance of the oocyte identity.

Publications

  • Mavrakis M, Rikhy R, Lilly MA, Lippincott-Schwartz J. Fluorescence imaging techniques for studying Drosophila development. Curr Protoc Cell Biol 2008 4:Unit 4.18.
  • Narbonne-Reveau K, Lilly MA. The Cyclin-dependent kinase inhibitor Dacapo promotes genomic stability during premeiotic S phase. Mol Biol Cell 2009 20:1960-1969.
  • Narbonne-Reveau K, Senger S, Pal M, Herr A, Asano M, Richardson HE, Deak P, Lilly MA. APC/CFzr/Cdh1 promotes cell cycle progression during the Drosophila endocycle. Development 2008 135:1451-1461.
  • Ullah Z, Lee CY, Lilly MA, Depamphilis ML. Developmentally programmed endoreduplication in animals. Cell Cycle 2009 8:1501-1509.

Collaborators

  • Maki Asano, PhD, Duke University Medical Center, Durham, NC
  • Péter Deák, PhD, Institute of Biochemistry, Szeged, Hungary
  • Helena Richardson, PhD, Peter MacCallum Cancer Center, Melbourne, Australia

Contact

For more information, email mlilly@helix.nih.gov or visit cbmp.nichd.nih.gov/uccr.

Top of Page