ISOLATION OF A HUMAN 11P112 LIVER TUMOR SUPPRESSOR GENE

许多研究表明，一个或多个基因的失活或缺失， 肿瘤抑制基因可能代表了肿瘤发生的早期和/或必要步骤， 肝脏的肿瘤性转化。 此外，比较 染色体定位研究表明， 在人类和啮齿类动物中可能有一个共同的分子机制， 涉及类似肿瘤抑制基因的失活。 基于 通过这些观察，我们认为染色体转移研究 利用人染色体和大鼠肝肿瘤细胞系可以 有助于人类基因的简单定位、鉴定和克隆 负责肝脏肿瘤抑制的基因。 为此我们 建立并表征了功能识别模型 人肿瘤抑制基因在大鼠肝肿瘤细胞系中的表达。 人类 11号染色体，它与人类的发病机制有关， 肝肿瘤，被引入高度侵袭性的大鼠肝上皮细胞中， 使用微细胞介导的染色体转移的肿瘤细胞系 法 我们已经证明，人类11号染色体抑制了 一些大鼠肝肿瘤细胞系的致瘤潜力，提供 肝肿瘤抑制基因存在的直接证据， 这个人类染色体 此外，我们还建立了一个微型卫星- 基于肝脏肿瘤抑制区的图谱，定位基因 人11p11.2-p12的950-kb片段，并形成了 候选肿瘤抑制基因的定位克隆。 研究 本提案中所载的内容旨在确定、分离和描述 人11p11.2-p12肝肿瘤抑制基因。 这个的目标 建议是（i）建立跨越肝肿瘤的PAC重叠群 人11p11.2-p12的抑制区，（ii）利用分离的PAC 克隆在构建一个完整的物理地图， 染色体区域，（iii）分离候选肝肿瘤抑制基因 基于与对应于该区域的PAC克隆的互补性， (iv)肝肿瘤抑制基因序列分析 分析，和（v）确定候选基因表达的能力， 转染率肝肿瘤细胞系中的肿瘤抑制活性。

Numerous studies suggest that inactivation or deletion of one or more tumor suppressor genes may represent early and/or necessary steps in the neoplastic transformation of liver. Furthermore, comparative chromosomal mapping studies suggest that the development of liver tumors in humans and rodents may share a common molecular mechanism that involves inactivation of analogous tumor suppressor genes. Based upon these observations, we have suggested that chromosome transfer studies utilizing human chromosomes and rat liver tumor cell lines may facilitate the facile localization, identification, and cloning of human genes responsible for tumor suppression in liver. To this end, we have established and characterized a model for the functional identification of human tumor suppressor genes in rat liver tumor cell lines. Human chromosome 11, which has been implicated in the pathogenesis of human liver tumors, was introduced into highly aggressive rat liver epithelial tumor cell lines using a microcell-mediated chromosome transfer technique. We have demonstrated that human chromosome 11 suppresses the tumorigenic potential of some rat liver tumor cell lines, providing direct evidence for the presence of a liver tumor suppressor gene on this human chromosome. Further, we have established a microsatellite- based map of the liver tumor suppressor region, localizing the gene(s) to a 950-kb segment of human 11p11.2-p12 and forming the basis for positional cloning of candidate tumor suppressor genes. The studies contained in this proposal seek to identify, isolate, and characterize the human 11p11.2-p12 liver tumor suppressor gene. The goals of this proposal are to (i) establish a PAC contig spanning the liver tumor suppressor region of human 11p11.2-p12, (ii) utilize the isolated PAC clones in the construction of a complete physical map of this chromosomal region, (iii) isolate candidate liver tumor suppressor genes based upon complementarity to PAC clones corresponding to this region, (iv) characterize candidate liver tumor suppressor genes by DNA sequence analysis, and (v) determine the ability of candidate genes to express tumor suppressor activity in transfected rate liver tumor cell lines.