MOLECULAR MECHANISM OF TAT INDUCED ANGIOGENESIS

TAT诱导血管生成的分子机制

• 批准号: 6056590
• 负责人: CINDY Anne MORRIS
• 金额: $ 21.45万
• 依托单位: LSU HEALTH SCIENCES CENTER
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-15 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6056590
• 关键词: BCL2 gene /protein; HeLa cells; RNase protection assay; SDS polyacrylamide gel electrophoresis; angiogenesis; angiogenesis factor; athymic mouse; cell adhesion molecules; cell differentiation; cell migration; cell proliferation; cyclin dependent kinase; cytokine; enzyme activity; fibroblast growth factor; human immunodeficiency virus 1; immunoprecipitation; laboratory mouse; northern blottings; nuclear runoff assay; transcription factor; vascular endothelium; virus protein; western blottings

DESCRIPTION (Adapted from the applicant's description) Human immunodeficiency virus type-1 (HIV-1)Tat is a potent transcriptional transactivator of both viral and cellular gene expression that possesses angiogenic properties. Tat cooperates with basic fibroblast growth factor(bFGF) in increasing the development of angiogenic lesions of Kaposi's sarcoma (KS) in HIV-1 infected individuals. Preliminary data suggest that this effect of Tat is triggered in vivo by inflammatory cytokines (IC)present in KS, including interleukin-1 beta (IL-1 beta), tumor necrosis factor-alpha (TNF-alpha) and gamma-interferon (gamma-IFN). Although both bFGF and vascular endothelial growth factor (VEGF) are expressed by KS cells in vivo and in vitro and IC upregulate the expression of both of these angiogenic factors, Tat exerts its angiogenic effects only with bFGF. The angiogenic process is complex, involving vascular endothelial cell growth, adhesion, migration, invasion and differentiation. To dissect the mechanism(s) for Tat-mediated angiogenic effects during each of these steps, in vitro and in vivo assays in which Tat, Tat mutants and Tat peptides are active have been developed. The central hypothesis of this proposal is that specific functional domains of Tat modulate proliferative, migratory and/or morphogenic effects on primary vascular endothelial cells during angiogenesis through a bFGF-mediated pathway. Toward this end, preliminary studies suggest that the ability of Tat to promote angiogenesis in synergy with bFGF is mediated, in part, by the selective interaction of the RGD domain of Tat with specific integrins (alpha5 beta1 and alphav Beta3) that are induced by bFGF. Through this interaction, the RGD domain of Tat promotes the locomotion and adhesion of KS and cytokine-activated primary vascular endothelial cells. In addition, the basic region of Tat retrieves extracellular bound bFGF into a soluble form that mediates cellular growth induced by Tat. Thus, Tat, a key regulatory protein of HIV-1 that can be released extracellularly from HIV-1-infected cells, upregulates angiogenesis, in part, by mimicking the functions of extracellular matrix proteins. The angiogenic effects of intracellular Tat are not known and determining this is a major focus of this proposal. The aims of the studies proposed herein are to determine whether (1) the transcriptional activating function of Tat, along with the RGD and basic region domains of Tat, plays a role during angiogenesis, (2) an endothelial cell-derived cyclin-dependent kinase, TAK/pTEF-b, activity that is essential for Tat transactivation, is induced through activation of primary human vascular endothelial cells by inflammatory cytokines or angiogenic factors that mediate Tat-enhanced angiogenesis and (3) Tat transactivates Bcl-2 gene expression in vascular endothelial cells stimulated to undergo angiogenesis, an effect that may involve the cellular Tat cofactor, TAK/pTEF-b.

描述(改编自申请人的描述) 人类免疫缺陷病毒1型(HIV-1)Tat是一种有效的转录因子 病毒和细胞基因表达的反式激活因子 血管生成特性。TAT配合碱性成纤维细胞生长 碱性成纤维细胞生长因子在促进Kaposi‘s血管新生病变发展中的作用 HIV-1感染者肉瘤(KS)。初步数据显示， TAT的这种作用是在体内由炎症细胞因子触发的 (IC)存在于KS，包括白介素1β(IL-1β)、肿瘤坏死 因子-α(肿瘤坏死因子-α)和干扰素(γ-干扰素)。虽然两者都是 KS细胞表达碱性成纤维细胞生长因子和血管内皮生长因子 体内、体外和IC均可上调这两种基因的表达 血管生成因子，TAT只有在碱性成纤维细胞生长因子的作用下才发挥其血管生成作用。这个 血管生成过程复杂，涉及血管内皮细胞生长， 黏附、迁移、侵袭和分化。要仔细分析 这些步骤中TAT介导的血管生成效应的机制(S)， TAT、TAT突变体和TAT多肽的体外和体内检测 Active已经开发出来了。这项提议的中心假设是 TAT的特定功能结构域调节增殖、迁移和/或 原代血管内皮细胞的形态发生作用 通过碱性成纤维细胞生长因子介导的血管生成。为此，初步 研究表明，TAT在促进血管生成方面具有协同作用 与碱性成纤维细胞生长因子的结合部分是由RGD的选择性相互作用所介导的 具有特定整合素(α5β1和αvβ3)的TAT结构域 均由碱性成纤维细胞生长因子诱导。通过这种相互作用，TAT的RGD结构域 促进KS和细胞因子激活的原代细胞的运动和黏附 血管内皮细胞。此外，TAT的基本区域检索 细胞外结合碱性成纤维细胞生长因子为可溶形式，可调节细胞生长 由TAT诱导。因此，Tat，HIV-1的关键调节蛋白，可以是 从感染HIV-1的细胞外释放，上调 血管生成，部分是通过模仿细胞外基质的功能实现的 蛋白质。细胞内TAT的血管生成作用尚不清楚， 确定这一点是这项提案的主要重点。这些研究的目的 在这里提出的是确定(1)转录激活 TAT的功能与TAT的RGD和基本区域结构域一起发挥作用 在血管生成中的作用，(2)内皮细胞衍生的细胞周期蛋白依赖 TAT反式激活所必需的激酶，即TAK/pTEF-b，是 通过激活原代人血管内皮细胞诱导 介导TAT增强的炎性细胞因子或血管生成因子 血管生成和(3)TAT反式激活血管内皮细胞中Bcl2基因的表达 刺激内皮细胞进行血管生成，这一效应可能 涉及细胞TAT辅因子TAK/pTEF-b。