ISOLATION OF A SECOND WILMS TUMOR SUPRESSOR GENE

人类肿瘤抑制基因的识别带来了新的见解 研究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抑制基因 使用分子标记或细胞遗传学。我们采取了相反的做法 通过使用一种名为肿瘤抑制的生物测试来绘制 功能性抑癌基因的单染色体定位 调职。通过这种方式，我们已经定位了一个肿瘤抑制基因 Wilms‘s瘤细胞系位于染色体11p15.5的两侧 匿名标记。我们初步绘制的这一区域的物理地图显示 该区域包含不到1000千个碱基(Kb)。基于相似性 在我们的功能研究和关于Rb和P53基因的研究之间， 我们认为该基因代表了另一个细胞周期控制基因。至 验证这一假设，我们将使用位置克隆技术来 确定功能性肿瘤抑制基因的候选基因。我们有 从这里收集了宇宙染色体和酵母人工染色体(YAC) 用于探测胎肾cdna文库的区域。为了对这个群进行分类， 我们将在Wilms的肿瘤细胞系中寻找具有异常的基因。 或者，我们将选择在基因中表达增加的基因 与亲本细胞系相关的微细胞杂交。终极的 这种肿瘤抑制基因的鉴定将取决于它的能力 抑制Wilms‘s肿瘤细胞系的致瘤性。一旦我们有了 分离出抑癌基因，我们将寻找序列同源性 与其他已知基因进行比较，以进一步描述其功能。之后 为基因产品开发免疫试剂，我们将 该基因在正常肾母细胞瘤细胞系中的表达 细胞和非致瘤微细胞杂交体。我们还将确定其 人胎儿发育过程中及不同组织中的表达模式 正常人体组织的类型。最后，我们将搜索异常情况 在Wilms‘s肿瘤患者的肿瘤样本中的这种基因 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。该抑癌基因的分离 将是第一个通过功能测试确定的这样的基因。这个 另一个细胞周期控制基因的出现将扩大我们的 了解肿瘤抑制基因的功能，可能会为 关于正常哺乳动物组织发育过程的线索。

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.