Nakayama Hironao, Sakaue Tomohisa, Maekawa Masashi, Fujisaki Ayako, Higashiyama Shigeki
Division of Cell Growth and Tumor Regulation, Proteo-Science Center, Ehime University, Ehime, Japan; Department of Biochemistry and Molecular Genetics, Ehime University Graduate School of Medicine, Toon, Shitsukawa, Ehime 791-0295, Japan; Department of Medical Science and Technology, Hiroshima International University, Higashi-hiroshima, Hiroshima 739-2695, Japan.
Division of Cell Growth and Tumor Regulation, Proteo-Science Center, Ehime University, Ehime, Japan; Department of Biochemistry and Molecular Genetics, Ehime University Graduate School of Medicine, Toon, Shitsukawa, Ehime 791-0295, Japan; Department of Cardiovascular and Thoracic Surgery, Ehime University Graduate School of Medicine, Toon, Shitsukawa, Ehime 791-0295, Japan.
Biochem Biophys Res Commun. 2018 Apr 30;499(1):17-23. doi: 10.1016/j.bbrc.2018.03.097. Epub 2018 Mar 21.
A disintegrin and metalloproteinase (ADAM) family are crucial enzymes for ectodomain shedding of multiple substrates and are involved in diverse biologic and pathologic processes. However, the molecular mechanism underlying substrate selectivity of ADAMs is poorly understood. In this study, we observed that disruption of actin polymerization by pharmacological inhibitors, latrunculin A (LatA) and cytochalasin D (CyD), induced ectodomain shedding of epidermal growth factor (EGF) family ligands. Induced shedding activity by LatA or CyD was suppressed by a metalloprotease inhibitor KB-R7785, indicating that ADAMs-mediated shedding is tightly controlled by actin cytoskeleton. We also investigated roles of cullin family, a component of cullin-RING based E3 ubiquitin ligases, in ectodomain shedding, since cullin family is implicated in the regulation of cytoskeletal dynamics. Knockdown of cullin 3 (Cul3) by a specific siRNA inhibited ectodomain shedding of amphiregulin (AREG), a member of EGF family, and responses were associated with activation of RhoA GTPase and induction of stress fiber formation. On the other hand, the RhoA inhibitor C3 transferase rescued AREG shedding reduced by Cul3 knockdown. These results describe a novel molecular mechanism of Cul3 to regulate AREG shedding by modulating cytoskeletal dynamics in a RhoA dependent manner.
解整合素金属蛋白酶（ADAM）家族是多种底物胞外域脱落的关键酶，参与多种生物学和病理过程。然而，对于ADAM底物选择性背后的分子机制，人们了解甚少。在本研究中，我们观察到，药理学抑制剂拉春库林A（LatA）和细胞松弛素D（CyD）破坏肌动蛋白聚合，会诱导表皮生长因子（EGF）家族配体的胞外域脱落。LatA或CyD诱导的脱落活性被金属蛋白酶抑制剂KB-R7785抑制，这表明ADAM介导的脱落受到肌动蛋白细胞骨架的严格控制。我们还研究了作为基于cullin-RING的E3泛素连接酶组分的cullin家族在胞外域脱落中的作用，因为cullin家族与细胞骨架动力学的调节有关。用特异性小干扰RNA敲低cullin 3（Cul3）可抑制EGF家族成员双调蛋白（AREG）的胞外域脱落，且这些反应与RhoA GTP酶的激活及应力纤维形成的诱导相关。另一方面，RhoA抑制剂C3转移酶挽救了因Cul3敲低而减少的AREG脱落。这些结果描述了Cul3通过以RhoA依赖的方式调节细胞骨架动力学来调控AREG脱落的一种新分子机制。
