171 Oral - Patterning and Morphogenesis II
Saturday April 09, 11:15 AM - 11:30 AM

Authors:
Zhi-Rong Ruan ¹; Adwait Amod Sathe ²; Chao Xing ²; Elizabeth Chen ¹

Affiliations:
1) Department of Molecular Biology, UT Southwestern Medical Center, Dallas, TX; 2) Eugene McDermott Center for Human Growth and Development, UT Southwestern Medical Center, Dallas, TX

Keywords:
j. muscle; a. core promoters and general transcription factors

Myoblast fusion, in which mononucleated myoblasts fuse to form multinucleated myotubes, is a critical step in skeletal muscle development and regeneration. In Drosophila, myoblast fusion is initiated by the interaction between muscle cell type-specific adhesion molecules. Here, we show that ecdysone signaling provides an extrinsic cue to regulate the onset of myoblast fusion by activating the expression of a cell adhesion molecule (CAM). Ecdysone is a steroid hormone best known for transcription regulation during pupal metamorphosis via binding to the ecdysone receptor (EcR). Surprisingly, we found that mutation in an ecdysteroidogenic enzyme, Spook (Spo), caused partial myoblast fusion defects in Drosophila embryos. Spo was expressed in amnioserosa, a major source for ecdysone production in early embryogenesis. The fusion defect in spo mutant was rescued by expressing spo in amnioserosa, but not in muscle cells, demonstrating a non-autonomous function of ecdysone at the tissue level. Since ecdysone is also provided maternally, we further disrupted ecdysone signaling by expressing a dominant negative form of EcR in muscle cells of spo mutant, which led to a severe fusion defect. Interestingly, the expression of Dumbfounded (Duf), a CAM required for the initiation of myoblast fusion, was significantly decreased in the spo;EcR^DN mutant. Conversely, overexpressing Duf in the spo;EcR^DN mutant significantly rescued the myoblast fusion defect, demonstrating that Duf is a major downstream target of ecdysone signaling in muscle cells. Indeed, we identified EcR-binding sites in the promoter of the duf gene and showed by in vitro luciferase assays that these sites mediated transcription activation by EcR. Furthermore, the EcR-binding sites are adjacent to the binding sites of a muscle-specific transcription factor, Twist. EcR and Twist synergistically activated duf transcription in vitro and in vivo. Taken together, these findings reveal an indispensable role for the extrinsic ecdysone signaling in regulating myoblast fusion and identify duf as one of the direct targets of ecdysone signaling in muscle cells. In addition, our studies demonstrate that the extrinsic ecdysone signaling coordinates with the intrinsic transcription factor Twist to synergistically activate the expression of duf to control the onset of myoblast fusion.