Project 3 Function of Kindlin-3 in blood and endothelial cells

摘要 这项建议的重点是整合素激活的机制，这是至关重要的基本细胞反应 对于血栓形成、炎症和血管炎等正常生理学和病理学是必需的。 功能障碍整合素在血管生成中起关键作用，或从现有的血管形成新的血管。 脉管系统有必要了解血管生成是如何调节的，以便创新的方法， 控制具有不同治疗需求的患者（即癌症、年龄相关性 黄斑变性和糖尿病视网膜病变）。我们的机械重点是金德林斯，一个 新的细胞内介体家族，其是所有细胞上的整联蛋白功能所需的。虽然我们的 以前和初步的研究表明，Kindlin在人类病理学中的重要性，许多Kindlin的 职能仍有待确定。因此，本项目的目标是进一步了解分子 Kindlin依赖整合素和独立整合素功能的机制和结构要求 在血管形成过程中的内皮细胞和骨髓细胞中。要检验的主要假设是，K3 是血管生成中内皮细胞和骨髓细胞功能所必需的， 功能依赖于整合素。提出了以下目标：目标1。为了确定K3- 内皮细胞和骨髓细胞中整联蛋白的依赖性调节在体内血管形成中起关键作用。 我们将通过以下方法检验K3-整联蛋白相互作用对体内血管生成调控至关重要的假设： 使用新产生的K3 KI敲入小鼠（表达不能与整联蛋白结合的K3突变体）。我们将 比较K3对内皮细胞和髓系细胞的作用。目标二。评估负责的机制 K3 KI小鼠的内皮细胞和髓样细胞功能受损，并用K3 和K2。确定EC中K3特异性功能的结构要求。目标3：以确定是否 以及K3在造血细胞中的功能在整合素活化过程中是如何调节的，重点是 K3磷酸化的调节作用。

Abstract This proposal focuses on the mechanisms of integrin activation, which is crucial for basic cellular responses essential for normal physiology and pathologies ranging from thrombosis, inflammation, and vascular dysfunction. Integrins play a key role in angiogenesis, or the formation of new blood vessels from existing vasculature. There is a need to understand how angiogenesis is regulated so that innovative approaches for the control of excessive vascular growth in patients with distinct therapeutic needs (i.e. cancer, age-related macular degeneration and retinopathy of diabetes) can be identified. Our mechanistic focus is on Kindlins, a novel family of intracellular mediators which are required for integrin functions on all cells. Although our previous and preliminary studies demonstrated importance of Kindlins in human pathologies, many Kindlins' functions remain to be identified. Thus, the goal of this project is to further understand the molecular mechanisms and structural requirements underlying integrin-dependent and independent functions of kindlins in endothelial and myeloid cells during vascularization processes. The main hypothesis to be tested is that K3 is required for functions of endothelial and myeloid cells in angiogenesis and many, but not all, of these functions are integrin-dependent. The following Aims are proposed: Aim 1. To determine whether K3- dependent regulation of integrins in endothelial and myeloid cells plays a critical role in vascularization in vivo. We will test the hypothesis that K3-integrin interaction is crucial for the regulation of angiogenesis in vivo by using newly generated K3KI knockin mice (expressing a mutant of K3 that is unable to bind to integrin). We will compare the role for K3 on endothelial to that on myeloid cells. Aim 2. To assess the mechanisms responsible for impaired function of endothelial and myeloid cells from K3KI mice, and perform rescue experiments with K3 and K2. To determine the structural requirement for K3-specific functions in EC. Aim 3. To determine whether and how K3 function in hematopoietic cells is regulated during integrin activation with an emphasis on the regulatory role of K3 phosphorylation.