Research and Professional Interests:

Cell-Cell Signaling In The Regulation Of Cell Cycle Programs And Oocyte Polarity In Drosophila

There are two major areas of research that are currently going on in our lab:

1. Oocyte Polarity, Drosophila Dystroglycan And Muscular Dystrophy

How polarity is established at a cellular level is one of the most fundamental and fascinating questions in cell and developmental biology. During development, many cell types undergo polarization to adopt forms that are highly adaptable to their specific functions. Disruption of cellular polarity frequently leads to abnormal cellular functions such as cell death, uncontrolled cell division, and irregular cell movement. Cytoskeletons and cell-cell and cell-substrate interactions are known to play important roles in cell polarization, but the mechanisms by which cellular polarity is established, modified, and maintained remain largely unknown.

The Drosophila oocyte is polarized along an anterior-posterior (AP) axis, which is also the basis for the AP body axis of the adult fly. The establishment of oocyte polarity requires a series of symmetry-breaking steps. Among these, cell-cell signaling between the oocyte and the follicle cells is critical. Two well established signaling pathways, the Epidermal Growth Factor Receptor (EGFR) pathway and the Notch pathway, both originating from the germline cells, are required for the differentiation of the follicle cells. Disruption of either pathway causes defects in oocyte polarity. It has been hypothesized that upon activation of the EGFR and Notch signaling, the posterior follicle cells (PFC) send an unidentified signal back to the oocyte, which re-orients the oocyte microtubules and, in turn, determines the polarity of the oocyte and the future embryo. The mechanism by which the forward signals (EGFR and Notch) and the mysterious back signal are connected is not clear.

Our recent studies have unveiled an emergent signaling pathway, the Dystroglycan pathway, in communicating the follicle cells to the oocyte. DG is a central component of the Dystrophin-glycoprotein complex (DGC) and plays an important role in maintaining normal muscle function. Disruption of this complex leads to various forms of muscular dystrophy in humans. The mammalian DG contains covalently linked single-transmembrane b -DG and highly glycosylated extracellular a -DG. At the cytoplasmic side, the proline-rich COOH terminus of b -DG binds directly to the WW domain of Dystrophin (Dys), whereas the a -DG binds to the Laminin G domains in Laminin-2, agrin, and perlecan with varying affinities. Thus, DG and Dys form a link to connect the ECM to the actin cytoskeleton.

Although DG is known to be required in a wide range of tissue and cell types, understanding of its exact cellular role is still vague. We will explore its role in intercellular communication in Drosophila. The lessons we learn in the model organism will be applicable to the mammalian system and will shed new light on the mechanism of its function in animal development and various pathological processes.



Figure 1. Dystroglycan is required in the germline for correct oocyte polarity during early oogenesis (A, B). (A) The wild-type oocyte has enriched actin staining (arrows), but the DG germ-line clone disrupts this enrichment (B). (From Deng et al., 2003)

2. Notch Signaling In Cell-Cycle Regulation

The Drosophila follicle-cell epithelium provides an excellent model system for the study of cell-cycle regulation in development and cell differentiation. During oogenesis, the follicle cells undergo three distinctive types of cell cycle programs: (1) the archetypal (normal) mitotic cycle; (2) the endoreplication cycle (also called the endocycle), in which cells duplicate their genomic DNA without cell division; and (3) amplification of specific genomic regions. These cell-cycle variants take place during different stages of oogenesis, and the switch from one variant to another is regulated by developmental signals.

The transition from the mitotic cycle to the endocycle is induced by Delta-Notch signaling during stage 6/7 of oogenesis. Components of the canonical Notch pathway, which include the ligand Delta, the gamma-secretase component presenilin, and the nuclear transducer Supressor of Hairless (Su(H)), are required in this process. Notch signaling down-regulates mitotic cyclins such as Cyclins A and B and the cdc25 homolog, String, during the transition.

In a recent study, we showed that Cut, a homeodomain protein, is required for follicle-cell proliferation and maintenance of an immature fate during early oogenesis. Its expression is down-regulated by Notch during the mitotic cycle/endocycle switch. This down-regulation is required for the switch from the mitotic cycle to the endocycle and cell differentiation and thus places Cut as one of the linkers between Notch signaling and cell-cycle regulators. Currently, we have found several more candidate genes that potentially function between the Notch pathway and the cell-cycle regulators through a mosaic screen. Further analysis of these genes will advance knowledge of cell-cycle regulation by developmental signals and the mechanisms by which Notch signaling is involved in regulating cell differentiation and cell proliferation. .



Figure 2. Switch of cell-cycle programs and the Cut expression pattern in follicle cells. (A) During Drosophila oogenesis, somatically derived follicle cells undergo two cell-cycle switches: (1) mitotic cycle to endocycle switch and (2) endocycle to gene-amplification switch. From the germarium (G) to stage (S) 6, follicle cells undergo unsynchronized mitotic cycles. During stages 7 to 10A, these cells go through three rounds of endoreplication and thereafter switch to a localized amplification pattern characteristic of chorion gene amplification. (B) Cut expression (shown in red) begins in follicle cells in region 2b of the germarium. It persists in all follicle cells, including the polar cells and the interfollicular stalk cells, until about stage 6 and diminishes afterwards, concurrent with the first cell-cycle switch. PH3 (shown in green) was used to mark the M phase of the mitotic cycle. Between stages 7 and 10A of oogenesis, Cut expression ceases in all follicle cells except the polar cells. At about stage 10B, Cut expression resumes in the columnar follicle cells that surround the oocyte (From Sun and Deng, 2005).

The Notch pathway has been implicated in a wide range of biological and pathological processes. In humans, defects in this pathway are implicated in multiple diseases, including leukemia, Alzheimer’s, and heart diseases. More recently, Notch has been shown to behave like a tumor-suppressor gene in mouse skin. Although much progress has been made toward understanding of the signaling process since its cloning in Drosophila 20 years ago, much remains to be learned about tuning of the signal in a developmental context and about its targets. The finding that Notch regulates a switch from normal mitotic cycle to endocycle links this important signaling to the intrinsic cell-cycle machinery, misregulation of which is frequently the cause of tumorigenesis.

Selected Publications:

Sun J, Deng WM. Hindsight mediates the role of notch in suppressing hedgehog signaling and cell proliferation. Dev Cell. 2007 Mar;12(3):431-42.

Schneider M, Khalil AA, Poulton J, Castillejo-Lopez C, Egger-Adam D, Wodarz A, Deng WM, Baumgartner S. (2006) Perlecan and Dystroglycan act at the basal side of the Drosophila follicular epithelium to maintain epithelial organization. Development. 133: 3805-15.

Poulton JS, Deng WM. 2006. Dystroglycan down-regulation links EGF receptor signaling and anterior-posterior polarity formation in the Drosophila oocyte. Proc Natl Acad Sci U S A. 103: 12775-12780.

Sun J., Deng W.M. 2005. Notch-dependent downregulation of the homeodomain gene cut is required for the mitotic cycle/endocycle switch and cell differentiation in Drosophila follicle cells. Development 132. 4299-4308.

Althauser C., Jordan K., Deng WM, Ruohola-Baker H. 2005. Fringe-dependent notch activation and tramtrack function are required for specification of the polar cells in Drosophila oogenesis. Developmental Dynamics 232: 1013-1020.

Schaeffer V., Althauser C., Shcherbata HR, Deng WM, Ruohola-Baker H. 2004. Notch-dependent Fizzy-related/Hec1/Cdh1 expression is required for the mitotic-to-endocycle transtition in Drosophila follicle cells. Current Biology 14(7): 630-6.

Deng, W.-M., M. Schneider, R. W. Frock, C. Castillejo-Lopez, S. Baumgartner, and H. Ruohola-Baker. 2003. Dystroglycan is required for polarizing the epithelial cells and the oocyte in Drosophila . Development 130: 173-184. (COVER PHOTOGRAPH).

Deng, W.-M., C. Althauser, and H. Ruohola-Baker. 2001. Notch-Delta signaling induces a transition from mitotic cell cycle to endocycle in Drosophila follicle cells . Development 128: 4737-4746. (COVER PHOTOGRAPH).

Deng, W.-M., and H. Ruohola-Baker. 2000. Laminin A is required for follicle cell-oocyte signaling that leads to establishment of the anterior-posterior axis in Drosophila . Current Biology 10: 683-686.

Jordan, K. C., N. J. Clegg, J. A. Blasi, A. M. Morimoto, J. Sen, D. Stein, H. McNeill, W.-M. Deng, M. Tworoger, and H. Ruohola-Baker. 2000. The homeobox gene mirror links EGF signaling to embryonic dorso-ventral axis formation through notch activation. Nature Genetics 24: 429-433.

Larkin, M. K.,* W. M. Deng,* K. Holder, M. Tworoger, N. Clegg, and H. Ruohola-Baker. 1999. Terminal follicle cell fate determination in Drosophila oogenesis. Development Genes & Evolution 209: 301-311. (*equal contribution)

Deng, W.-M., K. Leaper, and M. Bownes. 1999. A targeted gene silencing technique shows that Drosophila myosin VI is required for egg chamber and imaginal disc morphogenesis. Journal of Cell Science 112: 3677-3690.

Tzolovsky, G.,* W.-M. Deng,* T. Schlitt, and M. Bownes. 1999. The function of the broad complex during Drosophila melanogaster oogenesis. Genetics 153: 1371-1383. (*equal contribution)

Hicks, J. L., W.-M. Deng, A. D. Rogat, K. G. Miller, and M. Bownes. 1999. Class VI unconventional myosin is required for spermatogenesis in Drosophila. Molecular Biology of the Cell 10: 4341-4353.

Deng, W.-M., and M. Bownes. 1998. Patterning and morphogenesis of the follicle cell epithelium during Drosophila oogenesis. International Journal of Developmental Biology 42: 541-552.

Deng, W.-M., and M. Bownes. 1997. Two signalling pathways specify localised expression of the broad complex in Drosophila eggshell patterning and morphogenesis. Development 124: 4639-4647.

Deng, W.-M., D. Zhao, K. Rothwell, and M. Bownes. 1997. Analysis of P[gal4] insertion lines of Drosophila melanogaster as a route to identifying genes important in the follicle cells during oogenesis. Molecular Humam Reproduction 5: 101-110.

Tamori, Yoichiro
Tian, Ai-Guo
Wu, Xiaohui
Yu, Jianzhong

Graduate Students:

Ghaffar-Jabbari, Laila
Klusza, Stephen
Poulton, John
Sun, Jianjun

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