Identification of Receptor for Tumor Suppressor HIN-1

简介(由申请人提供)：建议的整体目标 研究目的是分离和鉴定HIN-1(High In 正常-1)。Hin-1是最近发现的一种可能的肿瘤抑制基因 SAGE(基因表达序列分析)用于比较基因表达谱 在正常和DCIS(导管原位癌，一种早期乳腺肿瘤)中 乳腺上皮细胞。Hin-1似乎是一种新的细胞因子， 在94%的人类乳腺癌中显著下调。这个 这项拟议研究的第一个具体目标是确定HIN-1的受体 使用一种表达克隆技术。生成的表达式库 Hin-1反应性乳腺上皮细胞将用碱性筛选 磷酸酶标记的HIN-1。拟议研究的具体目标2是(A) 通过执行以下操作确认HIN-1直接与分离的受体结合 共沉淀试验，以及(B)确定HIN-1与 通过全细胞结合分析分离受体。具体目标3是 目的：检测HIN-1受体在乳腺癌细胞系中的分布。 用实时定量聚合酶链式反应和荧光定量技术检测正常和DCIS乳腺上皮 原位杂交。一种HIN-1受体的鉴定与鉴定 开始表征HIN-1信号接收的第一步是必要的吗 和转导，这些研究可能进一步表明这种信号的损失 可能与乳腺肿瘤的进展有关。

DESCRIPTION (provided by applicant): The overall objective of the proposed research is to isolate and characterize a receptor for HIN-1 (High In Normal-1). HIN-1 is a putative tumor suppressor gene recently identified using SAGE (Serial Analysis of Gene Expression) to compare gene expression profiles in normal and DCIS (ductal carcinoma in-situ, an early-stage breast tumor) mammary epithelial cells. HIN-1 appears to be a novel cytokine that is significantly down-regulated in 94 percent of human breast carcinomas. The first specific aim of the proposed research is to identify a receptor for HIN-1 using an expression cloning technique. An expression library generated from HIN-1 responsive mammary epithelial cells will be screened with alkaline phosphatase-tagged HIN-1. Specific aim 2 of the proposed research is to (A) confirm that HIN-1 binds directly to the isolated receptor by performing a co-precipitation assay, and (B) determine the affinity of HIN-1 for the isolated receptor by performing whole cell binding analysis. Specific aim 3 is to determine distribution of the HIN-1 receptor in breast cancer cell lines and in normal versus DCIS mammary epithelium using real-time quantitative PCR and in-situ hybridization. Identification and characterization of a HIN-1 receptor is the first step necessary to begin characterization of HIN-1 signal reception and transduction, and these studies may further suggest how loss of this signal may contribute to breast tumor progression.