Release of FGF1 and the Pathology of Angiogenesis

FGF1 的释放和血管生成的病理学

• 批准号: 6750717
• 负责人: IGOR PRUDOVSKY
• 金额: $ 30.52万
• 依托单位: MAINEHEALTH
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-01-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6750717
• 关键词: angiogenesis; biological signal transduction; cell proliferation; exocytosis; fibroblast growth factor; intracellular transport; laboratory mouse; laboratory rabbit; protein structure function; protein transport; secretion; secretory protein; stress; synaptotagmin; vascular endothelium

The long-term goal of this laboratory has been to elucidate the mechanism by which fibroblast growth factor (FGF) 1 regulates the migration and proliferation of human endothelial cells in vitro and to apply this information to augment angiogenic responses during tissue and organ repair in vivo. While the recent application of the FGF prototypes for the generation of human collateral vessel growth in response to ischemic cardiovascular damage argues for the validity of this premise, structure-function studies with the members of the FGF gene family suggest that our understanding of FGF function is still primitive. Indeed, the function of two novel structural features within the FGF prototypes are not well understood and these include the presence of a functional nuclear/nucleolar localization signal(s) and the absence of a classical signal peptide sequence to direct its secretion through the conventional ER-Golgi apparatus-mediated pathway. Interestingly, these features are also found within members of the interleukin (IL)1 gene family which shares structural and crystallographic similarities with members of the FGF gene family. Since FGFs must gain access to the extracellular compartment to signal through their high affinity receptor tyrosine kinases in order to promote a biological response and undergo receptor-dependent nuclear/nucleolar translocation, we have utilized the resources from this award to define the mechanism by which FGF1 is released. Indeed, we have demonstrated during the current funding period that FGF1 but not FGF2 is released in response to biological stresses including heat shock, hypoxia, and serum deprivation in an apoptotic-independent manner. Structure- function analysis of the stress-induced FGF1 release pathway has demonstrated that FGF1 is exported into the extracellular compartment as a latent Cys30 FGF1 homodimer. The FGF1 homodimer is a component of a non-covalent high molecular weight complex which includes the extravesicular domain (p40) of the exocytotic trafficking-protein, p65 synaptotagmin (Syt1) and the annexin (Anx)2-binding protein, S100A13. Interestingly, like FGF1 and IL1alpha, these proteins also lack a classical signal peptide sequence and each is able to associate with acidic phospholipids including phosphatidylserine. In addition, we present preliminary data suggesting that the pathway responsible for the release of the mature but not the precursor form of the signal peptide-less pro-inflammatory cytokine, IL1alpha, is similar to the pathway utilized by FGF1 and that the precursor domain of IL1alpha acts as a dominant negative for the stress-induced release of FGF1. As a result, we request support to confirm and expand these results and propose to further define the mechanism of FGF1 release by (i) defining the structural prerequisites for the release of FGF1 and (ii) characterizing the cellular mechanisms by which these polypeptides facilitate FGF export. We suggest that these studies may not only provide new insight into the regulation of angiogenesis but may also enable access to novel pharmacologic targets for potential therapeutic management in man.

本实验室的长期目标是阐明成纤维细胞生长因子（FGF）1在体外调节人内皮细胞迁移和增殖的机制，并将此信息应用于体内组织和器官修复过程中增强血管生成反应。虽然最近应用的FGF原型的生成人侧支血管生长的缺血性心血管损伤的有效性，这一前提的论点，结构功能的研究与FGF基因家族的成员表明，我们的理解FGF的功能仍然是原始的。 事实上，FGF原型中两种新结构特征的功能尚未得到很好的理解，这些特征包括功能性核/核仁定位信号的存在和经典信号肽序列的缺失，以指导其通过常规ER-高尔基体介导的途径分泌。 有趣的是，这些功能也发现在白细胞介素（IL）1基因家族的成员，共享结构和晶体学的相似性与FGF基因家族的成员。 由于FGF必须进入细胞外区室，通过其高亲和力受体酪氨酸激酶发出信号，以促进生物反应并进行受体依赖性核/核仁易位，因此我们利用该奖项的资源来定义FGF 1释放的机制。 事实上，我们已经证明，在目前的资助期间，FGF 1，而不是FGF 2释放响应生物应激，包括热休克，缺氧，血清剥夺在一个独立的方式。 应激诱导的FGF 1释放途径的结构-功能分析已经证明FGF 1作为潜伏的Cys 30 FGF 1同源二聚体被输出到细胞外区室中。 FGF 1同源二聚体是非共价高分子量复合物的组分，该复合物包括胞吐运输蛋白p65突触结合蛋白（Syt 1）的囊外结构域（p40）和膜联蛋白（Anx）2结合蛋白S100 A13。 有趣的是，像FGF 1和IL 1 α一样，这些蛋白质也缺乏经典的信号肽序列，并且每种蛋白质都能够与酸性磷脂（包括磷脂酰丝氨酸）缔合。 此外，我们目前的初步数据表明，负责释放成熟的，但不是前体形式的信号肽少的促炎细胞因子，IL 1 α，是类似的途径利用FGF 1和IL 1 α的前体结构域作为一个显性负应力诱导的释放FGF 1。 因此，我们请求支持以确认和扩展这些结果，并建议通过（i）定义FGF 1释放的结构先决条件和（ii）表征这些多肽促进FGF输出的细胞机制来进一步定义FGF 1释放的机制。 我们认为，这些研究不仅可以提供新的见解调节血管生成，但也可能使获得新的药理学目标的潜在治疗管理的人。