Department of Medicine, University of California, San Diego, La Jolla, California, USA.
Department of Medicine, University of California, San Diego, La Jolla, California, USA.
J Biol Chem. 2021 Jan-Jun;296:100675. doi: 10.1016/j.jbc.2021.100675. Epub 2021 Apr 16.
Interaction of talin with the cytoplasmic tails of integrin β triggers integrin activation, leading to an increase of integrin affinity/avidity for extracellular ligands. In talin KO mice, loss of talin interaction with platelet integrin αIIbβ3 causes a severe hemostatic defect, and loss of talin interaction with endothelial cell integrin αVβ3 affects angiogenesis. In normal cells, talin is autoinhibited and localized in the cytoplasm. Here, we used an optogenetic platform to assess whether recruitment of full-length talin to the plasma membrane was sufficient to induce integrin activation. A dimerization module (Arabidopsis cryptochrome 2 fused to the N terminus of talin; N-terminal of cryptochrome-interacting basic helix-loop-helix domain ended with a CAAX box protein [C: cysteine; A: aliphatic amino acid; X: any C-terminal amino acid]) responsive to 450 nm (blue) light was inserted into Chinese hamster ovary cells and endothelial cells also expressing αIIbβ3 or αVβ3, respectively. Thus, exposure of the cells to blue light caused a rapid and reversible recruitment of Arabidopsis cryptochrome 2-talin to the N-terminal of cryptochrome-interacting basic helix-loop-helix domain ended with a CAAX box protein [C: cysteine; A: aliphatic amino acid; X: any C-terminal amino acid]-decorated plasma membrane. This resulted in β3 integrin activation in both cell types, as well as increasing migration of the endothelial cells. However, membrane recruitment of talin was not sufficient for integrin activation, as membrane-associated Ras-related protein 1 (Rap1)-GTP was also required. Moreover, talin mutations that interfered with its direct binding to Rap1 abrogated β3 integrin activation. Altogether, these results define a role for the plasma membrane recruitment of talin in β3 integrin activation, and they suggest a nuanced sequence of events thereafter involving Rap1-GTP.
与整合素β的胞质尾部相互作用的塔林触发整合素激活，导致整合素对细胞外配体的亲和力/亲合力增加。在塔林 KO 小鼠中，塔林与血小板整合素αIIbβ3 的相互作用丧失导致严重的止血缺陷，而塔林与内皮细胞整合素αVβ3 的相互作用丧失影响血管生成。在正常细胞中，塔林是自动抑制的，并定位于细胞质中。在这里，我们使用光遗传学平台来评估全长塔林募集到质膜是否足以诱导整合素激活。一个二聚化模块（拟南芥隐花色素 2 融合到塔林的 N 端；隐花色素相互作用的碱性螺旋-环-螺旋结构域的 N 端末端带有 CAAX 框蛋白 [C：半胱氨酸；A：脂肪族氨基酸；X：任何 C 末端氨基酸]）对 450nm（蓝色）光有反应，被插入中国仓鼠卵巢细胞和分别表达αIIbβ3 或αVβ3 的内皮细胞中。因此，细胞暴露于蓝光会导致拟南芥隐花色素 2-塔林快速且可逆地募集到隐花色素相互作用的碱性螺旋-环-螺旋结构域的 N 端末端带有 CAAX 框蛋白 [C：半胱氨酸；A：脂肪族氨基酸；X：任何 C 末端氨基酸]-装饰的质膜。这导致两种细胞类型中的β3 整合素激活，以及内皮细胞的迁移增加。然而，塔林的膜募集对于整合素激活来说并不足够，因为膜相关的 Ras 相关蛋白 1（Rap1）-GTP 也是必需的。此外，干扰塔林与其直接结合的 Rap1 的突变会阻断β3 整合素的激活。总之，这些结果定义了塔林在β3 整合素激活中的质膜募集作用，并表明随后涉及 Rap1-GTP 的事件有一个细微的顺序。
