HDAC9 nuclear/cytoplasmic shuttling in pulmonary vascular endothelial barrier regulation

HDAC9核/细胞质穿梭在肺血管内皮屏障调节中的作用

基本信息

批准号：
10446078
负责人：
ALEXANDER D VERIN
金额：
$ 38.5万
依托单位：
AUGUSTA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-02 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10446078
关键词：
Acute Lung Injury Acute Respiratory Distress Syndrome Adherens Junction Affect Agonist Antibiotics Bacteria Bacterial Infections Bacterial Toxins Blood Vessels Cell Nucleus Clinical Complex Cytoplasm Cytoplasmic Protein Cytoskeletal Proteins Cytosol Data Deacetylation Development Endothelial Cells Endothelium Escherichia coli Functional disorder GAB1 gene Goals HDAC9 gene HGF gene Histone Deacetylase Human In Vitro Inflammation Inflammatory Response Intensive Care Investigation Knockout Mice Lead Link Lipopolysaccharides Lung Mediating Modeling Molecular Mus Nuclear Nuclear Export Nuclear Import Pathologic Patients Permeability Pharmacology Phosphorylation Phosphotransferases Property Protein Dephosphorylation Protein phosphatase Proteins Pulmonary Circulation Pulmonary Edema Regulation Respiratory distress Role Sepsis Signal Transduction Signaling Protein Source Syndrome Testing Transcription Repressor Transgenic Mice Vascular Endothelial Cell Vascular Endothelium Vascular Permeabilities analog beta catenin genetic corepressor improved in vivo lung microvascular endothelial cells mortality novel novel therapeutic intervention overexpression plakoglobin protective effect protein activation pulmonary function receptor

项目摘要

PROJECT SUMMARY Acute lung injury (ALI) is characterized by lung vascular endothelial (EC) barrier compromise resulting in pulmonary edema. Gram negative (G-) bacteria are the source of bacterial toxins such as lipopolysaccharides (LPS), which are potent triggers of ALI. Despite the use of potent antibiotics and aggressive intensive care support, the mortality of LPS-induced ALI remains high, primarily because the complex molecular mechanisms involved in EC barrier regulation are ill-defined. The Scientific Premise of this Project derives from our novel preliminary data indicating that the class IIa histone deacetylase, HDAC9, is upregulated in the lungs of patients with sepsis in concert with the inflammatory responses. Accordingly, the overexpression of HDAC9 in transgenic mice increases basal lung vascular permeability and exacerbates vascular barrier compromise induced by LPS. Conversely, deletion of HDAC9 gene improves lung function in LPS-induced murine ALI model. These data suggest pro-edemagenic role of HDAC9 expression in ALI. HDAC9 is nuclear-cytoplasmic protein and its cellular localization is regulated by phosphorylation. Phosphorylation leads to HDAC9 nuclear export thus inhibiting its activity in the nucleus and promoting interactions with extra-nuclear targets. We demonstrated that LPS increases HDAC9 phosphorylation accompanied by its nuclear export in EC. In contrary, de-phosphorylation of HDAC9 leads to nuclear import thus inhibiting its association with cytoplasmic proteins. We showed that HDAC9 interacts with protein phosphatase 2A (PP2A), which has a barrier-protective role in EC. Further, our novel data indicated that PP2A is activated by Gi-mediated EC barrier-protective agonists, HGF and the stable ATP analog, ATPγS, suggesting the link between Gi-mediated signaling and PP2A in EC barrier enhancement. While the mechanisms of Gi-mediated PP2A activation have not been described, our data suggest that they may include the activation of GAB1/Shp2-mediated signaling. Further, both HDAC9 and PP2A interact with AJ protein plakoglobin suggesting functional complex. However, whether the HDAC9-dependent EC barrier regulation involved HDAC9/PP2A/plakoglobin interaction and EC barrier-protective effect of PP2A is mediated through decreased HDAC9 phosphorylation is not yet established. Therefore, a primary goal of this Proposal will be to define a novel role for HDAC9/PP2A crosstalk in AJ-mediated EC barrier regulation. We hypothesize that the phosphorylation status of HDAC9 and its interaction with PP2A alter intracellular signaling to regulate the EC barrier via effect on AJ assembly. The SAs are: 1: To evaluate the role of HDAC9 phosphorylation/nuclear export in the LPS-induced adherens junctions-mediated endothelial barrier compromise in vitro and in vivo. 2: To determine the importance of PP2A-mediated HDAC9 dephosphorylation/nuclear import in the assembly of AJs and endothelial barrier strengthening in vitro and in vivo.

项目概要急性肺损伤（ALI）的特点是肺血管内皮（EC）屏障受损，导致肺水肿。革兰氏阴性（G-）细菌是脂多糖等细菌毒素的来源。 (LPS)，尽管使用了强效抗生素并采取了积极的护理，但它仍然是 ALI 的有效触发因素。支持，LPS 诱导的 ALI 死亡率仍然很高，主要是因为复杂的分子机制该项目的科学前提源自我们的小说。初步数据表明 IIa 类组蛋白脱乙酰酶 HDAC9 在患者肺部表达上调脓毒症与炎症反应相一致，HDAC9在转基因中过度表达。小鼠的基础肺血管通透性增加，LPS 诱导的血管屏障受损恶化。在离线状态下，HDAC9 基因的缺失可改善 LPS 诱导的小鼠 ALI 模型中的肺功能。表明 HDAC9 表达在 ALI 中具有促水肿作用。磷酸化会导致 HDAC9 核输出，从而抑制其定位。我们证明了LPS在细胞核中的活性并促进与核外靶标的相互作用。增加 HDAC9 的磷酸化并伴随其在 EC 中的核输出，相反，HDAC9 的去磷酸化。 HDAC9 导致核输入，从而抑制其与细胞质蛋白的关联。与蛋白磷酸酶 2A (PP2A) 相互作用，后者在 EC 中具有屏障保护作用。表明 PP2A 被 Gi 介导的 EC 屏障保护激动剂、HGF 和稳定的 ATP 类似物激活， ATPγS，表明 Gi 介导的信号传导与 PP2A 在 EC 屏障增强中的联系。 Gi 介导的 PP2A 激活机制尚未描述，我们的数据表明它们可能包括 GAB1/Shp2 介导的信号传导的激活此外，HDAC9 和 PP2A 都与 AJ 蛋白相互作用。然而，plakoglobin 提示功能复合物是否依赖 HDAC9 的 EC 屏障调节。涉及 HDAC9/PP2A/plakoglobin 相互作用，PP2A 的 EC 屏障保护作用是通过 HDAC9 磷酸化尚未确定，因此该提案的主要目标是减少。定义了 HDAC9/PP2A 串扰在 AJ 介导的 EC 屏障调节中的新作用。 HDAC9 的磷酸化状态及其与 PP2A 的相互作用改变细胞内信号传导以调节 EC 屏障对 AJ 组装的影响 SA 是： 1：评估 HDAC9 磷酸化/核输出的作用。在体外和体内 LPS 诱导的粘附连接介导的内皮屏障受损中 2：To。确定 PP2A 介导的 HDAC9 去磷酸化/核导入在 AJ 组装中的重要性以及体外和体内内皮屏障的强化。