COLON CELL DIFFERENTIATION--DCC ROLE IN MUC2 EXPRESSION

结肠细胞分化--DCC 在 MUC2 表达中的作用

• 批准号: 2390901
• 负责人: LEONARD H AUGENLICHT
• 金额: $ 17.93万
• 依托单位: MONTEFIORE MEDICAL CENTER (BRONX, NY)
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-06-01 至 2001-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2390901
• 关键词: cell differentiation; colon neoplasms; gene expression; gene rearrangement; genetic enhancer element; genetic promoter element; goblet cells; human tissue; laboratory mouse; mucins; neoplasm /cancer genetics; phenotype; point mutation; protein biosynthesis; tissue /cell culture; transfection /expression vector; tumor suppressor genes

The DCC gene is a potential tumor suppressor gene whose deletion or inactivation plays a role in colonic tumor progression. Recent evidence suggests that it plays a role in colonic goblet cell differentiation and mucin production, consistent with its less can also be effected by inducers which interact with metabolic processes and signal transduction pathways. Some of these inducers may be related to dietary components. We are in a unique position to dissect this link among DCC, mucin synthesis and cell differentiation since we have initiated detailed studies on the mechanisms of regulation of the MUC2 gene, the gene which encodes the principal colonic mucin peptide backbone. We have now cloned, mapped and partially sequenced approximately 90 kilobases of the locus on chromosome II encompassing the MUC2 gene and its promotor and enhancer, and have found this contig contains another mucin backbone gene, probably MUC5b, which is also expressed in colonic cells both in vivo and in vitro, that could be coordinately regulated with MUC2. We will investigate the interaction between DCC and MUC2 expression at the RNA and protein level, as well as the elaboration and secretion of mucus and growth parameters. Colonic carcinoma cell lines which have deleted at least one copy of DCC will be used as recipients of expression vectors which contain the entire wild-type DCC cDNA, or which encode specific extracellular and intracellular domains of the DCC protein, and the influence on constitutive and inducible MUC2 expression determined. The experiments will be extended to mucin synthesis and secretion, since we have demonstrated that expression of the gene does not necessarily lead to production and secretion of mature mucins. Further, different promotor constructs will be used to define the relationship between specific levels of DCC expression and cell and molecular phenotype. The promoter and enhancer for MUC2 will be dissected to delineate sequence elements and their boundaries necessary to establish responsiveness of MUC2 to DCC. Finally, we will investigate this relationship of mutations in the promotor of the MUC2 gene to the sub-class of colonic carcinomas having a mucinous phenotype. These experiments will provide detailed understanding of the interaction of a gene important in the differentiation and transformation of colonic epithelial cells with the principal hallmark of differentiation along one lineage. In addition, since mucinous colonic tumors in which MUC2 is deregulated exhibit a poorer prognosis than most common colorectal tumors, understanding this interaction may define prognostic factors which are not reflected in tumor histology or pathology.

DCC基因是一种潜在的抑癌基因，其缺失或 失活在结肠肿瘤进展中发挥作用。 最近的证据 表明它在结肠杯状细胞分化中发挥作用 粘蛋白的产生与其较少一致，也可能受到诱导剂的影响 与代谢过程和信号转导途径相互作用。 其中一些诱导剂可能与饮食成分有关。 我们处于独特的地位来剖析 DCC、粘蛋白合成之间的这种联系 和细胞分化，因为我们已经开始了详细的研究 MUC2 基因的调节机制，该基因编码 主要结肠粘蛋白肽骨架。 我们现在已经克隆、映射并 对染色体上大约 90 KB 的基因座进行了部分测序 II 涵盖MUC2基因及其启动子和增强子，并发现 这个重叠群包含另一个粘蛋白主干基因，可能是MUC5b，它是 在体内和体外的结肠细胞中也表达，这可能是 与 MUC2 协调调节。 我们将研究 DCC 和 MUC2 表达之间的相互作用 RNA 和蛋白质水平，以及粘液的加工和分泌 和生长参数。 已删除的结肠癌细胞系 至少一份 DCC 将用作表达载体的接受者 包含完整的野生型 DCC cDNA，或编码特定的 DCC 蛋白的胞外和胞内结构域，以及 确定对组成型和诱导型 MUC2 表达的影响。 这 实验将扩展到粘蛋白的合成和分泌，因为我们 已经证明该基因的表达并不一定会导致 成熟粘蛋白的产生和分泌。 此外，不同的启动子 构造将用于定义特定级别之间的关系 DCC 表达以及细胞和分子表型的影响。 发起人及 MUC2 的增强子将被解剖以描绘序列元件和 它们的边界是建立 MUC2 对 DCC 反应性所必需的。 最后，我们将研究启动子突变的这种关系 将 MUC2 基因归为具有粘液性的结肠癌亚类 表型。 这些实验将提供对相互作用的详细理解 结肠分化和转化中重要的基因 具有沿一方向分化的主要特征的上皮细胞 血统。 此外，由于MUC2在粘液性结肠肿瘤中 与最常见的结直肠肿瘤相比，放松管制的预后较差， 了解这种相互作用可以定义预后因素，但这些因素不是 反映在肿瘤的组织学或病理学上。