ABSTRACT
Drosophila matrix metalloproteinase 2 (MMP2) is specifically expressed in posterior follicle cells of stage-14 egg chambers (mature follicles) and is crucial for the breakdown of the follicular wall during ovulation, a process that is highly conserved from flies to mammals. The factors that regulate spatiotemporal expression of MMP2 in follicle cells remain unknown. Here, we demonstrate crucial roles for the ETS-family transcriptional activator Pointed (Pnt) and its endogenous repressor Yan in the regulation of MMP2 expression. We found that Pnt is expressed in posterior follicle cells and overlaps with MMP2 expression in mature follicles. Genetic analysis demonstrated that pnt is both required and sufficient for MMP2 expression in follicle cells. In addition, Yan was temporally upregulated in stage-13 follicle cells to fine-tune Pnt activity and MMP2 expression. Furthermore, we identified a 1.1â kb core enhancer that is responsible for the spatiotemporal expression of MMP2 and contains multiple pnt/yan binding motifs. Mutation of pnt/yan binding sites significantly impaired the Mmp2 enhancer activity. Our data reveal a mechanism of transcriptional regulation of Mmp2 expression in Drosophila ovulation, which could be conserved in other biological systems.
Subject(s)
Drosophila Proteins , Drosophila , Animals , Female , Drosophila/metabolism , Matrix Metalloproteinase 2/genetics , Matrix Metalloproteinase 2/metabolism , Proto-Oncogene Proteins c-ets/genetics , Proto-Oncogene Proteins c-ets/metabolism , DNA-Binding Proteins/genetics , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Signal Transduction/physiology , Ovulation/genetics , Mammals/metabolism , Nerve Tissue Proteins/metabolism , Proto-Oncogene Proteins/metabolism , Transcription Factors/geneticsABSTRACT
Across species, ovulation is a process induced by a myriad of signaling cascades that ultimately leads to the release of encapsulated oocytes from follicles. Follicles first need to mature and gain ovulatory competency before ovulation; however, the signaling pathways regulating follicle maturation are incompletely understood in Drosophila and other species. Our previous work has shown that the bHLH-PAS transcription factor Single-minded (Sim) plays important roles in follicle maturation downstream of the nuclear receptor Ftz-f1 in Drosophila. Here, we demonstrate that Tango (Tgo), another bHLH-PAS protein, acts as a co-factor of Sim to promote follicle cell differentiation from stages 10 to 12. In addition, we discover that re-upregulation of Sim in stage-14 follicle cells is also essential to promote ovulatory competency by upregulating octopamine receptor in mushroom body (OAMB), matrix metalloproteinase 2 (Mmp2) and NADPH oxidase (NOX), either independently of or in conjunction with the zinc-finger protein Hindsight (Hnt). All these factors are crucial for successful ovulation. Together, our work indicates that the transcriptional complex Sim:Tgo plays multiple roles in late-stage follicle cells to promote follicle maturation and ovulation.
Subject(s)
Drosophila Proteins , Matrix Metalloproteinase 2 , Animals , Female , Basic Helix-Loop-Helix Transcription Factors/genetics , Basic Helix-Loop-Helix Transcription Factors/metabolism , Drosophila/metabolism , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Matrix Metalloproteinase 2/metabolism , Oogenesis/genetics , Ovulation/geneticsABSTRACT
Ovulation is a critical reproductive process by which a mature oocyte is released from the ovary for fertilization. This process requires the coordination of multiple cellular and molecular events including the spatiotemporal breakdown of the follicle wall. Recent work has shown that ovulation in Drosophila utilizes conserved cellular processes and molecular pathways as in mammals. Thus, Drosophila ovulation can serve as a good model to decipher the fundamental mechanisms of ovulation utilized across species. In past decades, several methods have been developed to study Drosophila ovulation, but all of them have drawbacks. This chapter offers a strategy and detailed protocols for performing and analyzing the necessary assays to evaluate the ovulation process in Drosophila.
