Regulation of B1 Integrin Glycosylation by RAS

描述：(申请人提供)整合素黏附受体调节许多 重要的细胞过程包括细胞增殖、细胞 迁移/侵袭、细胞分化和锚定依赖性生长。 这些区域中任何一个区域的整合素功能改变都可以促进肿瘤的发生 和/或转移，因此，密集的调查集中在 了解这些受体的调节。提交的初步研究 在这项提案中描述了一种新的整合素调节机制。它是 结果表明，致癌ras基因表达导致B1整合素获得5-6倍的 更丰富的a2-6连接唾液酸残基。一种指导酶的酶 这种连锁ST6Ga1 I在肿瘤组织中上调，并增加了ST6Ga1 I 与细胞侵袭性有关。相应地，ST6Ga1 i的增加 长期以来，活动一直被认为与癌症的进展有关。尽管如此 发现，该酶靶向的特定分子物种尚未 已确认身份。提出了一个中心假设，即致癌的ras增加了 ST6Ga1 I唾液酸基转移酶的表达和/或功能，并且这 然后，酶作用于B1整合素，增加a2-6唾液酸的数量 残留物。反过来，增加31整合素的2-6唾液酸化导致 导致功能改变的构象变化。广泛的、长期的 这项拟议研究的目标是证明变异糖基化 代表了B1整合素调节的重要机制，而且， 致癌ras诱导的一种变异糖型表达有助于 肿瘤的发生和/或转移。这些目标将通过以下方式实现：1) 确定ras介导的B1整合素碳水化合物结构和 ST6Ga1 I活性(目标1)以阐明ras改变的机制 整合素糖基化、N-连接的碳水化合物结构和位置 糖基化将被鉴定，ST6Ga1 I的活性/表达将 被评估。2)确定糖基化改变在 修改整合素功能(目标2)的整合素A2-6唾液酸化水平将是 直接改变和整合素的功能将被检查。此外，细胞 将使用可诱导的RAS结构来确定是否表达 糖型改变与功能改变在时间上是相关的。3) 确定131整合素糖基化改变是否与癌症有关 进展期(目标3)过表达ST6Ga1 I的细胞将被检查 非锚定生长、迁移/侵袭与裸体肿瘤的形成 老鼠。此外，还将检测胰腺肿瘤标本中 一种变种的B1糖型。

DESCRIPTION: (provided by applicant) Integrin adhesion receptors regulate many important cellular processes including cell proliferation, cell migration/invasion, cell differentiation and anchorage-dependent growth. Altered integrin function in any of these areas can promote tumorigenesis and/or metastasis, and hence, intense investigation has centered on understanding the regulation of these receptors. Preliminary studies presented in this proposal describe a novel mechanism for integrin regulation. It is shown that expression of oncogenic ras cause Bl integrins to acquire a 5-6-fold greater abundance of a2-6 linked sialic acid residues. The enzyme that directs this linkage, ST6Ga1 I, is upregulated in tumor tissues, and increased ST6Ga1 I is associated with cell invasiveness. Accordingly, increases in ST6Ga1 I activity have long been implicated in cancer progression. Despite these findings, the specific molecular species targeted by this enzyme have not been identified. A central hypothesis is proposed that oncogenic ras increases the expression and/or function of the ST6Ga1 I sialyltransferase, and that this enzyme then acts on the, B1 integrin to increase the number of a2-6 sialic acid residues. In turn, increased a2-6 sialylation of the 31 integrin induces a conformational change that leads to altered function. The broad, long term goals of the proposed research are to demonstrate that variant glycosylation represents an important mechanism for B1 integrin regulation and further, that oncogenic ras-induced expression of a variant glycoform contributes to tumorigenesis and/or metastasis. These goals will be accomplished by: 1) Defining the ras-mediated changes in B1 integrin carbohydrate structure and in ST6Ga1 I activity (Aim 1) To elucidate the mechanism by which ras alters integrin glycosylation, the carbohydrate structures and sites of N-linked glycosylation will be identified, and the activity/expression of ST6Ga1 I will be evaluated. 2) Establishing that altered glycosylation plays a causal role in modifying integrin function (Aim 2) Levels of integrin a2-6 sialylation will be directly altered and integrin function will be examined. In addition, cells with an inducible ras construct will be used to determine whether expression of an altered glycoform is temporally correlated with altered function. 3) Determining whether altered 131 integrin glycosylation contributes to cancer progression (Aim 3) Cells that overexpress ST6Ga1 I will be examined for anchorage-independent growth, migration/invasion, and tumor formation in nude mice. In addition, pancreatic tumor specimens will be assayed for expression of a variant B1 glycoform.