Yang Chansik, Ohk Jiyeon, Lee Ji Yeun, Kim Eun Jin, Kim Jiyoon, Han Sangyeul, Park Dongeun, Jung Hosung, Kim Chungho
From the Department of Life Sciences, Korea University, Seoul, Republic of Korea (C.Y., J.Y.L., E.J.K., J.K., C.K.); School of Biological Sciences, Seoul National University, Seoul, Republic of Korea (C.Y., D.P.); Department of Anatomy, Brain Research Institute, and Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea (J.O., H.J.); and Center for Vascular Research, Institute for Basic Science, Daejeon, Korea (S.H.).
Arterioscler Thromb Vasc Biol. 2016 Jul;36(7):1406-16. doi: 10.1161/ATVBAHA.116.307619. Epub 2016 May 19.
Angiogenesis, the process of building complex vascular structures, begins with sprout formation on preexisting blood vessels, followed by extension of the vessels through proliferation and migration of endothelial cells. Based on the potential therapeutic benefits of preventing angiogenesis in pathological conditions, many studies have focused on the mechanisms of its initiation as well as control. However, how the extension of vessels is terminated remains obscure. Thus, we investigated the negative regulation mechanism.
We report that increased intracellular calcium can induce dephosphorylation of the endothelial receptor tyrosine kinase Tie2. The calcium-mediated dephosphorylation was found to be dependent on Tie2-calmodulin interaction. The Tyr1113 residue in the C-terminal end loop of the Tie2 kinase domain was mapped and found to be required for this interaction. Moreover, mutation of this residue into Phe impaired both the Tie2-calmodulin interaction and calcium-mediated Tie2 dephosphorylation. Furthermore, expressing a mutant Tie2 incapable of binding to calmodulin or inhibiting calmodulin function in vivo causes unchecked growth of the vasculature in Xenopus. Specifically, knockdown of Tie2 in Xenopus embryo retarded the sprouting and extension of intersomitic veins. Although human Tie2 expression in the Tie2-deficient animals almost completely rescued the retardation, the Tie2(Y1113F) mutant caused overgrowth of intersomitic veins with strikingly complex and excessive branching patterns.
We propose that the calcium/calmodulin-dependent negative regulation of Tie2 can be used as an inhibitory signal for vessel growth and branching to build proper vessel architecture during embryonic development.
血管生成是构建复杂血管结构的过程，始于在已有血管上形成芽，随后通过内皮细胞的增殖和迁移使血管延伸。基于在病理状态下阻止血管生成的潜在治疗益处，许多研究聚焦于其起始及调控机制。然而，血管延伸如何终止仍不清楚。因此，我们研究了负调控机制。
我们报告细胞内钙增加可诱导内皮受体酪氨酸激酶Tie2去磷酸化。发现钙介导的去磷酸化依赖于Tie2-钙调蛋白相互作用。绘制了Tie2激酶结构域C末端环中的Tyr1113残基图谱，发现该相互作用需要此残基。此外，将该残基突变为苯丙氨酸会损害Tie2-钙调蛋白相互作用和钙介导的Tie2去磷酸化。此外，在体内表达不能与钙调蛋白结合或抑制钙调蛋白功能的突变型Tie2会导致非洲爪蟾血管系统不受控制地生长。具体而言，在非洲爪蟾胚胎中敲低Tie2会延迟体节间静脉的芽生和延伸。尽管在Tie2缺陷动物中表达人Tie2几乎完全挽救了这种延迟，但Tie2(Y1113F)突变体导致体节间静脉过度生长，具有极其复杂和过度的分支模式。
我们提出，钙/钙调蛋白对Tie2的负调控可作为一种抑制信号，在胚胎发育过程中抑制血管生长和分支，以构建合适的血管结构。
