ISOLATION OF A SECOND WILMS TUMOR SUPRESSOR GENE

第二个 Wilms 肿瘤抑制基因的分离

• 批准号: 2104868
• 负责人: Bernard E. Weissman
• 金额: $ 22.33万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-06-01 至 1997-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2104868
• 关键词: Wilms' tumor; artificial chromosomes; athymic mouse; breast neoplasms; cell cycle proteins; chromosome deletion; gene expression; genetic library; genetic markers; human fetus tissue; human tissue; hybrid cells; molecular cloning; molecular oncology; neoplasm /cancer genetics; plasmids; protein sequence; rhabdomyosarcoma; tumor suppressor genes; western blottings

The identification of human tumor suppressor genes has led to new insights into the mechanisms of human cancer development. Most of the known tumor suppressor genes were isolated by determining their chromosomal location using molecular markers or cytogenetics. We have taken the opposite approach by using a biological assay, tumor suppression, to map the locations of functional tumor suppressor genes via monochromosome transfer. In this manner, we have mapped a tumor suppressor gene for a Wilms' tumor cell line to a region of chromosome 11p15.5 flanked by two anonymous markers. Our preliminary physical map of this area suggests that this region contains less than 1000 kilobases (kb). Based on similarities between our functional studies and those concerning the RB and p53 genes, we propose that this gene represents another cell cycle control gene. To test this hypothesis, we will use positional cloning techniques to identify candidates for the functional tumor suppressor gene. We have gathered both cosmids and yeast artificial chromosomes (YACs) from this area for probing a fetal kidney cDNA library. To sort through this group, we will look for genes with abnormalities in the Wilms' tumor cell line. Alternatively, we will chose genes which show increased expression in the microcell hybrids relative to the parental cell line. The ultimate identification of this tumor suppressor gene will depend on its ability to suppress tumorigenicity in the Wilms' tumor cell line. Once we have isolated the tumor suppressor gene, we will search for sequence homologies with other known genes to further delineate its functions. After developing immunological reagents for the gene product, we will characterize expression of the gene in Wilms' tumor cell lines, normal cells and non-tumorigenic microcell hybrids. We will also determine its patterns of expression in developing human fetal material and in different types of normal human tissues. Finally, we will search for abnormalities in this gene in tumor samples from Wilms' tumor patients as well as patients with other malignant and/or genetic diseases which map to this same region of chromosome 11. The isolation of this tumor suppressor gene will represent the first such gene identified by a functional assay. The availability of another cell cycle control gene would broaden our understanding of tumor suppressor gene functions and may provide important clues about the process of normal mammalian tissue development.

人类肿瘤抑制基因的鉴定带来了新的见解 人类癌症发展的机制。大多数已知的肿瘤 通过确定抑制基因的染色体位置来分离它们 使用分子标记或细胞遗传学。我们采取了相反的做法 方法通过使用生物测定，肿瘤抑制，映射 功能性肿瘤抑制基因的单染色体定位 转移以这种方式，我们已经定位了一个肿瘤抑制基因， Wilms肿瘤细胞系的染色体11p15.5区域，两侧是两个 匿名标记。我们对这一地区的初步物理地图显示， 该区域含有少于1000个内切酶（kb）。基于相似性 在我们的功能研究和RB和p53基因的研究之间， 我们认为该基因代表另一个细胞周期控制基因。 到 为了验证这一假设，我们将使用定位克隆技术， 鉴定功能性肿瘤抑制基因的候选者。我们有 收集了cosmopolysaccharides和酵母人工染色体（YACs）， 用于探测胎儿肾cDNA文库的区域。为了对这组人进行分类， 我们将在肾母细胞瘤细胞系中寻找异常基因。 或者，我们将选择在细胞中表达增加的基因。 微细胞杂交相对于亲本细胞系。最终 这种肿瘤抑制基因鉴定将取决于其 抑制肾母细胞瘤细胞系的致瘤性。一旦我们 分离肿瘤抑制基因后，我们将寻找序列同源性 与其他已知基因进行比对以进一步阐明其功能。后 为基因产物开发免疫试剂，我们将 表征Wilms肿瘤细胞系中基因的表达，正常 细胞和非致瘤性微细胞杂交体。我们还将确定其 在发育中的人胎儿材料和不同的人胚胎材料中的表达模式 正常人体组织的类型。最后，我们将寻找异常 在肾母细胞瘤患者的肿瘤样本中， 患有其他恶性和/或遗传性疾病的患者， 11号染色体的同一区域。这种肿瘤抑制基因的分离 将代表通过功能测定鉴定的第一个这样的基因。的 另一个细胞周期控制基因的可用性将扩大我们的研究范围。 了解肿瘤抑制基因的功能，并可能提供重要的 正常哺乳动物组织发育过程的线索。