NOVEL ZINC FINGER PROTEIN THAT INHIBITS TNF CYTOTOXICITY

肿瘤坏死因子等细胞因子对血管内皮细胞有多向作用 各种各样的细胞和组织。这些变化尤其是 在内皮细胞中，肿瘤坏死因子被证明能诱导黏附 对白细胞有促进作用，提高膜促凝血活性。更改 这一性质很可能是促炎作用的核心 肿瘤坏死因子。为了阐明肿瘤坏死因子改变的分子机制 内皮细胞功能，我们利用差异斑块杂交来 确定肿瘤坏死因子即刻早期反应基因。有七个这样的基因是 已确认身份。DNA测序显示，两个编码的已知细胞因子诱导了 基因；内皮细胞白细胞黏附分子-1与中性粒细胞趋化 因素。一个编码核原癌基因；c-jun，而另一个编码 一种新发现的单核细胞特异性趋化因子 与内皮相关。一种单核细胞趋化因子的生产 细胞因子激活的内皮细胞对理解 血管壁在疾病状态中的作用，如动脉粥样硬化和 也可能在一定程度上解释了肿瘤坏死因子的间接血管生成活性。 值得注意的是，根据数据判断，其他三个cDNA完全是新的 对部分DNA序列进行库搜索。重要的是，其中之一(A20) 包含七个锌指，这是某一类 转录因子。为了更好地理解单核细胞的作用 化学诱导剂和血管壁病状态下的新的cDNA， 血管生成和炎症我们建议进行以下研究： 目标1：(I)克隆编码新转录本的全长cDNA和 完成他们的DNA测序。具体目标2：(一)进行细菌融合 单核细胞趋化蛋白和新的cDNA 兔免疫和抗体产生。具体目标3：使用 单特异性多克隆抗血清的特异性2，定位 单核细胞趋化因子和新抗原：a)体外，使用 细胞因子处理内皮细胞和其他细胞。这将允许亚细胞 新基因产物的本地化。B)在体内，处于炎症状态 例如伤口愈合和牛皮癣。对于单核细胞趋化剂来说 对于检查动脉粥样硬化组织将特别重要。此外， 将进行原位杂交研究，以确定哪些细胞 在被研究的组织中，实际上正在转录感兴趣的基因。

Cytokines such as tumor necrosis factor (TNF) exert pleiotropic effects on a wide variety of cells and tissues. These alterations are particularly profound in endothelial cells where TNF has been shown to induce adherence for leukocytes and to increase membrane procoagulant activity. Changes of this nature are likely to be central to the pro-inflammatory effects of TNF. In order to elucidate molecular mechanisms by which TNF alters endothelial cell function, we utilized differential plaque hybridization to identify TNF immediate early response genes. SEVEN such genes were identified. DNA sequencing revealed that two encoded known cytokine induced genes; endothelial leukocyte adhesion molecule-1 and neutrophil chemotactic factor. One encoded the nuclear protooncogene; c-jun, while another encoded a recently described monocyte specific chemotactic factor NOT previously associated with endothelium. The production of a monocyte chemotaxin by cytokine activated endothelium has important implications for understanding the role of the vessel wall in disease states such as atherosclerosis and may also in part explain the indirect angiogenic activity of TNF. Significantly, the other three cDNAs are completely novel as judged by data bank searches of partial DNA sequences. Importantly, one of these (A20) contains seven zinc fingers which is characteristic of a certain class of transcriptional factors. To better understand the role of the monocyte chemoattractant and the novel cDNAs in vessel wall disease states, angiogenesis and inflammation we propose the following studies: Specific Aim 1: (i) Clone full length cDNAs encoding the novel transcripts and complete their DNA sequencing. Specific Aim 2: (i) Make bacterial fusion protein for both the monocyte chemoattractant and the novel cDNAs for rabbit immunization and antibody production. Specific Aim 3: Using the monospecific polyclonal antiserum raised in Specific Aim 2, localize both the monocyte chemoattractant and the novel antigens: a) In Vitro, using cytokine treated endothelial and other cells. This will allow subcellular localization of the novel gene products. b) In vivo, in inflammatory states such as wound healing and psoriasis. For the monocyte chemoattractant it will be particularly important to examine atherosclerotic tissue. Further, in situ hybridization studies will be carried out to identify which cells in the tissue under study are actually transcribing the gene of interest.