Fachbereich Medizin, Department for Molecular Cell Physiology, Institute for Physiology and Pathophysiology, Philipps-Universität Marburg, Emil-Mannkopff-Str. 2, 35037 Marburg, Germany.
Fachbereich Biologie, Department for Developmental Biology, Philipps-Universität Marburg, Karl-von-Frisch-Str. 8, 35043 Marburg, Germany.
Biol Open. 2024 Jan 15;13(1). doi: 10.1242/bio.058666. Epub 2024 Jan 5.
Guanine nucleotide exchange factors (GEF) of the BRAG subfamily activate small Arf GTPases, which are pivotal regulators of intracellular membrane traffic and actin dynamics. Consequently, BRAG proteins have been implicated to regulate the surface levels of adhesive and signaling receptors. However, not much is known about the mechanism leading to the regulation of these surface proteins. In this study, we found that the Drosophila BRAG GEF Schizo interacts physically with the Abl-interactor (Abi). schizo mutants display severe defects in myoblast fusion during syncytial muscle formation and show increased amounts of the cell adhesion protein N-cadherin. We demonstrate that the schizo myoblast fusion phenotype can be rescued by the expression of the Schizo GEF (Sec7) and membrane-binding (pleckstrin homology) domain. Furthermore, the expression of the Sec7-PH domain in a wild-type background decreases the amounts of N-cadherin and impairs myoblast fusion. These findings support the notion that the Sec7-PH domain serves as a constitutive-active form of Schizo. Using a yeast-two hybrid assay, we show that the SH3 domain of Abi interacts with the N-terminal region of Schizo. This region is also able to bind to the cytodomain of the cell adhesion molecule N-cadherin. To shed light on the function of Schizo and Abi in N-cadherin removal, we employed epistasis experiments in different developmental contexts of Drosophila. These studies point towards a new model for the regulation of Schizo. We propose that the binding of Abi to the N-terminal part of Schizo antagonizes Schizo function to inhibit N-cadherin removal.
鸟嘌呤核苷酸交换因子（GEF）的 BRAG 亚家族激活小的 Arf GTPases，这是细胞内膜运输和肌动蛋白动力学的关键调节因子。因此，BRAG 蛋白被认为调节粘附和信号受体的表面水平。然而，对于导致这些表面蛋白调节的机制知之甚少。在这项研究中，我们发现果蝇 BRAG GEF Schizo 与 Abl 相互作用蛋白（Abi）在物理上相互作用。schizo 突变体在合胞肌肉形成过程中肌母细胞融合严重缺陷，并显示出细胞粘附蛋白 N-钙粘蛋白的含量增加。我们证明，schizo 肌母细胞融合表型可以通过 Schizo GEF（Sec7）和膜结合（pleckstrin 同源）结构域的表达来挽救。此外，Sec7-PH 结构域在野生型背景下的表达降低了 N-钙粘蛋白的含量并损害了肌母细胞融合。这些发现支持 Sec7-PH 结构域作为 Schizo 的组成活性形式的观点。使用酵母双杂交测定，我们表明 Abi 的 SH3 结构域与 Schizo 的 N 端区域相互作用。该区域还能够与细胞粘附分子 N-钙粘蛋白的胞质结构域结合。为了阐明 Abi 在 N-钙粘蛋白去除中的作用，我们在果蝇的不同发育背景下进行了上位性实验。这些研究为 Schizo 的调控提供了一个新的模型。我们提出，Abi 与 Schizo N 端部分的结合拮抗了 Schizo 功能，抑制了 N-钙粘蛋白的去除。
