MOLECULAR MECHANISM OF TAT INDUCED ANGIOGENESIS

描述（根据申请人的描述改编） 人类免疫缺陷病毒1型（HIV-1）达特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。 达特与碱性成纤维细胞生长协同作用 因子（bFGF）增加卡波西血管生成性病变的发展 肉瘤（KS）在HIV-1感染的个人。 初步数据显示， 达特的这种作用在体内由炎性细胞因子触发 （IC）存在于KS中，包括白细胞介素-1 β（IL-1 β）、肿瘤坏死 因子-α（TNF-α）和γ-干扰素（γ-IFN）。 虽然两 KS细胞表达bFGF和血管内皮生长因子（VEGF 在体内和体外，IC上调这两种表达 血管生成因子，达特发挥其血管生成作用，只有与bFGF。 的 血管生成过程是复杂的，涉及血管内皮细胞生长， 粘附、迁移、侵袭和分化。 解剖 在这些步骤中的每一个中Tat介导的血管生成作用的机制， 体外和体内测定，其中达特、达特突变体和达特肽 积极发展。 这一提议的核心假设是， 达特特异性功能结构域调节增殖、迁移和/或 对原代血管内皮细胞形态发生的影响 血管生成通过bFGF介导的途径。 为此，初步 研究表明达特协同促进血管生成的能力 与bFGF的相互作用部分是由RGD的选择性相互作用介导的。 达特结构域与特异性整联蛋白（α 5 β 1和α v β 3）， 是由bFGF诱导的。 通过这种相互作用，达特的RGD结构域 促进KS的运动和粘附，并促进酪氨酸激活的初级 血管内皮细胞 此外，达特的基本区域检索 将细胞外结合的bFGF转化为介导细胞生长的可溶形式 由达特引起的。因此，达特是HIV-1的一种关键调节蛋白， 从HIV-1感染的细胞中细胞外释放， 血管生成，部分通过模拟细胞外基质的功能， proteins. 细胞内达特的血管生成作用尚不清楚， 确定这一点是本提案的主要重点。 研究的目的 本文提出的是确定（1）转录激活是否 达特的功能与达特的RGD和碱性区域结构域沿着， 血管生成过程中的作用，（2）内皮细胞衍生的细胞周期蛋白依赖性 激酶TAK/pTEF-b活性是达特反式激活所必需的， 通过激活原代人血管内皮细胞诱导， 炎症细胞因子或血管生成因子介导Tat增强 （3）达特反式激活血管内皮细胞Bcl-2基因表达 刺激内皮细胞进行血管生成，这种作用可能 涉及细胞达特辅因子TAK/pTEF-b。

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.