FUNCTIONAL STUDY OF A CARCINOMA ASSOCIATED MUCIN MUC1

我们的目标是建立MUC1粘蛋白在肿瘤发生和转移中所起作用的分子基础。MUC1是一种细胞相关的粘蛋白糖蛋白，在90%以上的癌症和转移性病变中高度过度表达。我们发现，缺乏MUC1的小鼠的自发性乳腺肿瘤在肿瘤生长速度和转移方面都有非常显著的降低。我们的假设是，MUC1是一种多功能蛋白质，其结构特征有助于其调节肿瘤的进展和转移。一个结构特征是棒状的，密集糖基化的胞外结构域，它调节肿瘤细胞的粘附性和它们对免疫效应细胞的脆弱性。第二个结构特征是酪氨酸磷酸化的细胞质尾部结构域，它与参与信号转导和细胞骨架重组的蛋白质相互作用。为了验证我们的假设，在目标1中，我们将在小鼠的乳腺中过表达人MUC1蛋白，直接模拟人类乳腺癌中MUC1过表达的表型。将对正常腺体的发育进行研究。这些小鼠将与发展为自发性乳腺癌的MMTV-MTag和MMTV-c-neu小鼠杂交，并将检测MUC1过度表达的影响。我们将监测肿瘤的进展和转移。2)为了研究MUC1域的功能意义，我们在正常腺体和肿瘤中缺失不同结构域(串联重复序列、胞外区和胞内区)的转基因小鼠中过表达MUC1区，以检测这些区域在肿瘤发生和发展中的功能。3)我们将利用酵母双杂交技术和免疫共沉淀实验鉴定与MUC1相互作用的蛋白质，并表征MUC1与参与信号转导和细胞骨架组织的细胞质蛋白相互作用的分子细节和后果。计划中的实验将加深我们对MUC1如何通过其棒状的胞外结构域以及通过细胞质结构域与参与信号转导的蛋白质相互作用来调控肿瘤进展和转移的理解。

Our objective is to establish the molecular basis for the contribution of the MUC1 mucin to tumorigenesis and metastasis. MUC1 is a cell associated mucin glycoprotein that is highly overexpressed in greater than 90 percent of carcinomas and metastatic lesions. We showed that spontaneous mammary gland tumors in mice lacking Muc1 had highly significant decreases in tumor growth rate and metastasis. Our hypothesis is that MUC1 is a multi-functional protein with structural features that contribute to its ability to modulate tumor progression and metastasis. One structural feature is the rodlike, densely glycosylated extracellular domain that modulates the adhesiveness of tumor cells and their vulnerability to immune effector cells. The second structural feature is the tyrosine phosphorylated cytoplasmic tail domain that interacts with proteins involved in signal transduction and cytoskeletal reorganization. To test our hypothesis, in aim 1 we will overexpress the human MUC1 protein in mammary glands of mice, directly mimicking the phenotype of MUC1 overexpression in human breast cancer. Development in the normal glands will be studied. These mice will be crossed with MMTV-MTag and MMTV-c-neu mice that develop spontaneous breast cancer and the bitransgenics will be examined for the effects of MUC1 overexpression. We will monitor tumor progression and metastasis. 2) To examine the functional significance of the MUC1 domains, we will overexpress MUC1 in transgenic mice with various domains deleted (tandem repeats, extracellular, and cytoplasmic domains) in normal glands and tumors to test the functions of these regions in tumorigenesis and development. 3) We will identify proteins interacting with MUC1 using the yeast two-hybrid technique and coimmunoprecipitation experiments and characterize the molecular details and consequences of MUC1 interactions with cytoplasmic proteins involved in signal transduction and cytoskeletal organization. The planned experiments will add to our understanding of how MUC1 modulates tumor progression and metastasis by virtue of its rodlike extracellular domains as well as through interactions of the cytoplasmic domain with proteins that are involved in signal transduction.