TRANSFORMATION OF CELLS BY THE V-REL ONCOGENE

REV-T是一种禽类逆转录病毒，专门在早期转化为 体内和体外的淋巴样细胞。REV-T癌基因为v-T。 Rel；v-Rel编码转化蛋白p59v-Rel。V-REL为 来源于正常的细胞基因c-rel，与 果蝇背部基因。我们对这个机制很感兴趣 V-Rel通过什么途径转化淋巴样细胞，以及为什么 不会转化成纤维细胞。使用链接器插入和 活体突变我们打算分离出v-rel突变体 对淋巴细胞的转化温度敏感。 这些突变体将被用于实验，以确定 如果p59v-rel是或与蛋白激酶活性相关 重要的是它的变换功能。如果激酶活性发生变化 对于v-rel变换函数WE似乎并不重要 将尝试确定一种相关的生化功能 P59v-Rel使用从细菌细胞中纯化的蛋白表达 V-Rel蛋白。我们还将确定磷酸化的位置 以及p59v中第二核导向函数的恒等式- REL，以及这些序列对于转换的重要性 功能使用定点突变。禽类蛋白p59v- 雷尼替丁对小鼠成纤维细胞有毒性。我们将确定V-Rel 负责这种毒性的序列，以及是否无毒 禽类Rel蛋白可转化小鼠淋巴样细胞。如果不是，我们 将尝试分离小鼠c-rel基因克隆，并在体外通过 我们将尝试创造一种显性的小鼠亲属 转化基因。使用DNA转移技术，我们将 试图从淋巴细胞中分离出基因，这将使 成纤维细胞的转化。最后，通过背靠背的方式 我们将确定逆转录病毒载体中的杂交基因是否 这两个基因之间的序列同源性也反映了 两种蛋白质之间的功能同源性。建议数 研究旨在加深我们对逆转录病毒的理解 转化过程，并可能与淋巴相关 肿瘤的发展和其他正常的发育过程 将军。

Rev-T is an avian retrovirus that specifically transforms early lymphoid cells in vivo and in vitro. The oncogene of Rev-T is v- rel; v-rel encodes the transforming protein p59v-rel. v-rel is derived from a normal cellular gene c-rel, and is highly related to the Drosophila dorsal gene. We are interested in the mechanism by which v-rel transforms lymphoid cells, and the reason why it does not transform fibroblast cells. Using linker insertion and in vivo mutagenesis we intend to isolate v-rel mutants that are temperature-sensitive for transformation of lymphoid cells. These mutants will be used in experiments designed to determine if p59v-rel is, or is asssociated with, a protein kinase activity important for its transforming function. If kinase activity does not seem to be important for the v-rel transforming function we will attempt to determine a pertinent biochemical function of p59v-rel using protein purified from bacterial cells expressing the v-rel protein. We will also determine the sites of phosphorylation and the identity of a second nuclear directing function in p59v- rel, and the importance of these sequences for the transforming function using site-directed mutagenesis. The avian protein p59v- rel is toxic in murine fibroblasts. We will determine the v-rel sequences responsible for this toxicity, and whether a non-toxic avian rel protein can transform murine lymphoid cells. If not, we will attempt to isolate a murine c-rel cDNA clone, and by in vitro mutagenesis we will attempt to create a dominant murine rel transforming gene. Using DNA transfer techniques, we will attempt to isolate genes from lymphoid cells that will allow transformation of fibroblast cells. Finally, by making rel-dorsal hybrid genes in retroviral vectors we will determine whether the sequence homology between these two genes also reflects a functional homology between the two proteins. The proposed studies are intended to further our understanding of retroviral transforming processes, and may have relevance to lymphoid tumor development and other normal developmental processes in general.