DISTRIBUTION AND FUNCTIONAL ASPECTS OF TGF-BETA-3

TGF-BETA-3 的分布和功能方面

• 批准号: 3193650
• 负责人: Leslie Ina Gold
• 金额: $ 14.02万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-06-15 至 1994-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3193650
• 关键词: carcinoma; cell bank /registry; chemical structure function; collagen; enzyme inhibitors; enzyme linked immunosorbent assay; extracellular matrix; fibronectins; high performance liquid chromatography; human tissue; immunofluorescence technique; lung neoplasms; neoplastic growth; plasminogen activator; protein sequence; protein tyrosine kinase; radioimmunoassay; transforming growth factors; umbilical cord; western blottings

A novel growth inhibitor of the TGF-beta-family has been cloned and sequenced from human placental cDNA; the new form is designated TGF-beta-3. TGF-betas are secreted as inactive precursor molecules, that are subsequently proteolytically cleaved to yield biologically active mature dimers of 25 KDa. The precursor regions of TGF-beta-1 and TGF-beta-3 contain an RGD sequence. TGF-betas influence the proliferation and differentiation processes of cells, and specifically demonstrate potent growth inhibitory activity on many cell types, especially carcinomas. Preliminary experiments showed that TGF-beta-3 exhibited at least 10 fold greater growth inhibitory activity on cells, including a human lung carcinoma. Recently, different functions, partially mediated through specific receptors have been demonstrated for TGF-beta-1 and TGF-beta-2. Since TGF-beta-3 is a newly discovered protein its distribution, functions, and receptor binding characteristics are unknown. Anti- sera will be raised to synthetic peptides of both the precursor and mature forms of TGF-beta-3. Non-cross-reacting anti-peptide IgG will be used analytically to screen normal and neoplastic cells and tissues for TGF-beta-3 by immunofluorescence. Immunoprecipitation and immunoaffinity chromatographic techniques will be employed to quantitate and purify mature TGF-beta-3 and the precursor form of TGF-beta-1 and TGF-beta-3 from cells and tissues. Purified TGF- beta-3 will be used to quantitate its specific activity in various TGF-beta specific functions and to identify and characterize its receptor binding entities on the cell surface. By stimulating the production of certain components of the extracellular matrix (ECM) such as fibronectin (Fn) and collagen, and by preventing ECM breakdown through the synthesis of anticatalytic proteins such as plasminogen activator inhibitor (PAI), TGF-beta appears to have an important effect on maintaining the integrity of the ECM. The influence of TGF-beta-3 on Fn, collagen, and PAI production will be assessed. The possibility that the inactive TGF-beta-3 precursor molecule binds to the Fn receptor through its RGD sequence, creating a negative feedback mechanism, will be examined. Blocking the binding of fibronectin to its receptor would cause decreased adherence of cells to the ECM. The Fn receptor is phosphorylated on tyrosines in transformed but not normal cells. The ability of TGF-beta-3 to ameliorate transformation related properties by preventing tyrosine phosphorylation of the Fn receptor will be evaluated. The significance of these studies in understanding control mechanisms of tumor cell proliferation and metastasis may elucidate concepts for future therapeutics in two important areas of cancer research.

一种新的TGF-β家族生长抑制剂已被克隆， 从人类胎盘cDNA测序;新的形式被命名为 TGF-β-3 TGF-β作为非活性前体分泌 分子，其随后被蛋白水解裂解以产生 生物活性成熟二聚体25 kDa。 前体区域 TGF-β-1和TGF-β-3中含有RGD序列。 TGF-β 影响细胞的增殖和分化过程， 并特别显示出有效的生长抑制活性， 许多细胞类型，尤其是癌。 初步实验 显示TGF-β-3的生长至少比对照组高10倍， 对细胞的抑制活性，包括人肺癌。 最近，不同的功能，部分介导的特定 已经证明了TGF-β-1和TGF-β-2的受体。 由于TGF-β-3是一种新发现的蛋白质，因此其分布， 功能和受体结合特性是未知的。 反 血清将被提高到合成肽的前体和 TGF-β 3的成熟形式。 非交叉反应抗肽IgG 将用于分析筛选正常和肿瘤细胞， TGF-β-3的免疫荧光。 免疫沉淀 免疫亲和层析技术将被用来 定量和纯化成熟TGF-β-3和TGF-β-3的前体形式， 来自细胞和组织的TGF-β-1和TGF-β-3。 纯化的TGF- β-3将用于定量其在各种环境中的比活性。 TGF-β特异性功能，并鉴定和表征其 细胞表面的受体结合实体。 通过刺激 细胞外基质（ECM）的某些成分的产生 如纤连蛋白（Fn）和胶原蛋白，并通过防止ECM 通过合成抗催化蛋白质，如 纤维蛋白溶酶原激活物抑制剂（派），TGF-β似乎具有 对保持ECM的完整性有重要作用。 的 TGF-β 3对Fn、胶原和派产生影响将 被评估。 失活的TGF-β-3的可能性 前体分子通过其RGD与Fn受体结合 序列，创造一个负反馈机制，将被审查。 阻断纤连蛋白与其受体的结合会导致 降低细胞对ECM的粘附。 Fn受体是 在转化细胞而不是正常细胞中酪氨酸磷酸化。 TGF-β-3改善转化相关性的能力 通过防止Fn的酪氨酸磷酸化的性质 将对受体进行评估。 这些研究的意义在于 了解肿瘤细胞增殖的控制机制， 转移可能阐明未来治疗的概念， 癌症研究的重要领域。