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

摘要 这项建议侧重于整合素激活的机制，整合素对基本的细胞反应至关重要。 对包括血栓、炎症和血管在内的正常生理和病理来说是必不可少的 功能障碍。整合素在血管生成中起关键作用，即从现有血管形成新血管。 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 控制有不同治疗需求的患者(如癌症、年龄相关的患者)的过度血管生长 糖尿病的黄斑变性和视网膜病变)可以被识别。我们的机械论重点是Kindlins，一个 所有细胞上的整合素功能所需的新型细胞内介质家族。虽然我们的 先前和初步的研究表明Kindlins在人类病理中的重要性，许多Kindlins的 功能还有待确定。因此，本项目的目标是进一步了解分子 Kindlins整合素依赖和独立功能的机制和结构要求 在血管形成过程中内皮细胞和髓样细胞中。需要检验的主要假设是K3 是内皮细胞和髓系细胞在血管生成中的功能所必需的，其中许多但不是全部 功能依赖于整合素。提出了以下目标：目标1.确定K3- 内皮细胞和髓系细胞中整合素的依赖调节在体内血管形成中起着关键作用。 我们将通过以下方式验证K3-整合素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K3KI敲打小鼠(表达无法与整合素结合的K3突变体)。我们会 比较K3在内皮细胞和髓系细胞上的作用。目标2.评估负责的机制 针对K3KI小鼠受损的内皮细胞和髓系细胞，进行K3的抢救实验 和K2。确定EC中K3特定功能的结构要求。目标3.确定是否 以及在整合素激活过程中K3如何在造血细胞中发挥作用，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.