Identification of Receptor for Tumor Suppressor HIN-1

描述（由申请人提供）：拟议的总体目标 研究是分离和表征HIN-1的受体（高In 正常-1）。HIN-1是一个假定的肿瘤抑制基因，最近发现， SAGE（基因表达系列分析）比较基因表达谱 在正常和DCIS（导管原位癌，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。的 这项研究的第一个具体目标是鉴定HIN-1的受体 使用表达克隆技术。一个表达式库， HIN-1反应性乳腺上皮细胞将用碱性磷酸酶筛选。 磷酸酶标记的HIN-1。本研究的具体目标2是（A） 确认HIN-1直接结合到分离的受体，通过进行 共沉淀测定，和（B）测定HIN-1对HIN-1的亲和力， 通过进行全细胞结合分析分离受体。具体目标三是 确定HIN-1受体在乳腺癌细胞系中的分布， 在正常与DCIS乳腺上皮中使用实时定量PCR， 原位杂交HIN-1受体的鉴定和表征 是开始表征HIN-1信号接收所必需的第一步 这些研究可能进一步表明，这种信号的丢失 可能会导致乳腺癌的进展。

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.