Department of Cardiovascular & Metabolic Sciences, Lerner Research Institute, Cleveland Clinic, Ohio, USA.
Department of Cardiovascular & Metabolic Sciences, Lerner Research Institute, Cleveland Clinic, Ohio, USA; Department of Biochemistry, School of Medicine, Case Western Reserve University, Cleveland, Ohio, USA.
J Biol Chem. 2021 Jan-Jun;296:100685. doi: 10.1016/j.jbc.2021.100685. Epub 2021 Apr 21.
Ras suppressor-1 (Rsu-1) is a leucine-rich repeat (LRR)-containing protein that is crucial for regulating cell adhesion and is involved in such physiological and pathological processes as focal adhesion assembly and tumor metastasis. Rsu-1 interacts with zinc-finger type multi-LIM domain-containing adaptor protein PINCH-1, known to be involved in the integrin-mediated consensus adhesome, but not with its highly homologous family member PINCH-2. However, the structural basis for and regulatory mechanisms of this specific interaction remain unclear. Here, we determined the crystal structures of Rsu-1 and its complex with the PINCH-1 LIM4-5 domains. Rsu-1 displays an arc-shaped solenoid architecture, with eight LRRs shielded by N- and C-terminal capping modules. We showed that the conserved concave surface of the Rsu-1 LRR domain binds and stabilizes the PINCH-1 LIM5 domain via salt bridge and hydrophobic interactions, while the C-terminal non-LIM region of PINCH-2 sterically disfavors Rsu-1 binding. We also showed that Rsu-1 can be assembled, via PINCH-1-binding, into a heteropentamer complex comprising Rsu-1, PINCH-1, ILK, Parvin, and Kindlin-2, which constitute a major consensus integrin adhesome crucial for focal adhesion assembly. Our mutagenesis and cell biological data emphasize the significance of the Rsu-1/PINCH-1 interaction in focal adhesion assembly and cell spreading, providing crucial molecular insights into Rsu-1-mediated cell adhesion with implications for disease development.
Ras 抑制因子-1（Rsu-1）是一种富含亮氨酸重复序列（LRR）的蛋白质，对调节细胞黏附至关重要，参与了诸如焦点黏附组装和肿瘤转移等生理和病理过程。Rsu-1 与锌指型多 LIM 结构域包含衔接蛋白 PINCH-1 相互作用，后者已知参与整合素介导的共识黏附体，但不与高度同源的家族成员 PINCH-2 相互作用。然而，这种特异性相互作用的结构基础和调控机制仍不清楚。在这里，我们确定了 Rsu-1 及其与 PINCH-1 LIM4-5 结构域复合物的晶体结构。Rsu-1 呈现出弧形螺旋结构，由 N 端和 C 端盖帽模块屏蔽的八个 LRR 组成。我们表明，Rsu-1 LRR 结构域的保守凹面通过盐桥和疏水相互作用结合并稳定 PINCH-1 LIM5 结构域，而 PINCH-2 的 C 端非 LIM 区域则阻碍了 Rsu-1 的结合。我们还表明，通过与 PINCH-1 的结合，Rsu-1 可以组装成一个异五聚体复合物，该复合物由 Rsu-1、PINCH-1、ILK、Parvin 和 Kindlin-2 组成，它们构成了一个主要的共识整合素黏附体，对焦点黏附组装至关重要。我们的突变和细胞生物学数据强调了 Rsu-1/PINCH-1 相互作用在焦点黏附组装和细胞扩展中的重要性，为 Rsu-1 介导的细胞黏附提供了关键的分子见解，并对疾病的发展具有重要意义。
