Pluskota Elzbieta, Szpak Dorota, Wang Yunmei, Qin Jun, Simon Daniel I, Plow Edward F, Bialkowska Katarzyna
Department of Cardiovascular and Metabolic Sciences, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA.
Case Cardiovascular Research Institute, Case Western Reserve University School of Medicine and Harrington Heart & Vascular Institute, University Hospitals Cleveland Medical Center, Cleveland, Ohio, USA.
Res Pract Thromb Haemost. 2025 Apr 17;9(3):102863. doi: 10.1016/j.rpth.2025.102863. eCollection 2025 Mar.
A single phosphorylation in kindlin-3 at S (human) or S (mouse) has been shown to regulate its function in a variety of cells However, whether this posttranslational modification of kindlin-3 plays a role in platelets and in hemostasis and thrombosis remains totally unknown.
To create a strain of mice expressing kindlin-3 that bears the SA substitution and utilize these mice to determine if kindlin-3 phosphorylation influences platelet responses, hemostasis, and thrombosis .
By clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 gene editing, we generated a mouse strain that expressed a kindlin-3 mutant bearing the SA substitution. SA kindlin-3 mice were born normally, and the platelet and red blood cells numbers were similar to their wild-type counterparts. Wild-type and SA kindlin-3 platelets showed similar expression of αβ integrin, kindlin-3, and talin.
Platelets isolated from SA kindlin-3 mice did not undergo agonist-induced kindlin-3 phosphorylation. Functional analysis revealed that SA kindlin-3 mice exhibited prolonged tail bleeding time, increased blood loss, and delayed thrombosis in the carotid artery injury model . Platelets isolated from SA kindlin-3 mice showed defective platelet function, including impaired integrin αβ activation, platelet aggregation, clot retraction, and adhesion.
Our observations demonstrate that kindlin-3 phosphorylation on S in mice is crucial for supporting activation of αβ integrin and αβ integrin-dependent platelet responses and consequently contributes to hemostasis and thrombosis .
已有研究表明，kindlin-3在人源S或鼠源S位点的单个磷酸化修饰可调节其在多种细胞中的功能。然而，这种kindlin-3的翻译后修饰在血小板以及止血和血栓形成过程中是否发挥作用仍完全未知。
构建表达携带S→A替换的kindlin-3的小鼠品系，并利用这些小鼠确定kindlin-3磷酸化是否影响血小板反应、止血和血栓形成。
通过成簇规律间隔短回文重复序列/CRISPR相关蛋白9基因编辑技术，我们构建了一个表达携带S→A替换的kindlin-3突变体的小鼠品系。S→A kindlin-3小鼠正常出生，其血小板和红细胞数量与野生型小鼠相似。野生型和S→A kindlin-3血小板的αⅡbβ3整合素、kindlin-3和踝蛋白表达相似。
从S→A kindlin-3小鼠分离的血小板未发生激动剂诱导的kindlin-3磷酸化。功能分析显示，S→A kindlin-3小鼠的尾部出血时间延长、失血量增加，并且在颈动脉损伤模型中血栓形成延迟。从S→A kindlin-3小鼠分离的血小板表现出血小板功能缺陷，包括整合素αⅡbβ3激活受损、血小板聚集、血块回缩和黏附受损。
我们的观察结果表明，小鼠中kindlin-3在S位点的磷酸化对于支持αⅡbβ3整合素激活以及αⅡbβ3整合素依赖性血小板反应至关重要，从而有助于止血和血栓形成。
