TRANSFORMING GENES OF AVIAN SARCOMA VIRUSES

本建议的目的是了解其性质， 禽肉瘤病毒转化基因的功能 （ASV）及其正常细胞对应部分原癌基因。 ASV UR 2的转化基因v-ros及其相应的 将研究细胞基因c-ros和原癌基因c-src。 v-ros的功能域将通过构建 各种位点特异性突变体以及ROS和SRC重组体。 ros蛋白的合成和亚细胞定位， 与潜在的细胞底物相互作用， 将在亲本UR 2和突变体之间比较能力。 c-ros基因在禽类和哺乳动物组织中的表达 各种发育阶段将在RNA和 蛋白质水平 将识别c-ros产品， 使用ROS特异性抗血清表征。 原位 将进行免疫荧光以鉴定特异性 组织中表达c-ros蛋白的细胞。 的cDNA克隆 将构建c-ros mRNA并测序，以阐明 完整的c-ros产物，其似乎模拟生长因子 受体分子 为了识别序列差异 在c-ros和v-ros之间， c-ros病毒和c-ros两个基因的转化潜力 将构建重组体，并与 它们的基因产物和致癌性。 已知鸡c-src指导4kb mRNA的合成， 其编码60，000道尔顿酪氨酸特异性蛋白激酶。 在肌肉中，当4kb的RNA缺失时，3 kb的c-src mRNA 缺少大部分激酶结构域反而被表达。 无论是4 将克隆3 kb和3 kb RNA并测序以阐明 它们的合成性质。 3的潜在蛋白质产物 kb RNA将通过使用抗肽的抗血清来鉴定 从核苷酸序列中解码出来。 这个src的作用- 与肌肉发育和功能相关的蛋白质将被 研究了

The objective of this proposal is to understand the nature and functions of the transforming genes of avian sarcoma viruses (ASVs) and their normal cellular counter-part proto-oncogenes. The transforming gene of ASV UR2, v-ros, its corresponding cellular gene, c-ros, and the proto-oncogene c-src will be studied. The functional domains of v-ros will be studied by constructing various site-specific mutants, and ros and src recombinants. Synthesis and subcellular location of the ros protein, its interaction with potential cellular substrates and transforming ability will be compared among parental UR2 and the mutants. The expression of c-ros gene in avian and mammalian tissues at various developmental stages will be examined at RNA and protein levels. The c-ros product will be identified and characterized using ros-specific antisera. In situ immunofluorescence will be performed to identify the specific cells in the tissue expressing the c-ros protein. The cDNA clone of c-ros mRNA will be constructed and sequenced to elucidate the complete c-ros product, which appears to mimic growth factor receptor molecules. To identify the sequence differences between c-ros and v-ros that may be responsible for the different transforming potential of the two genes, c-ros virus and c-ros recombinants will be constructed and compared with respect to their gene products and oncogenicity. Chicken c-src is known to direct the synthesis of a 4 kb mRNA which codes for a 60,000 dalton tyrosine-specific protein kinase. In muscle, while the 4 kb RNA is absent, a 3 kb c-src mRNA lacking most the kinase domain is expressed instead. Both the 4 kb and the 3 kb RNAs will be cloned and sequence to elucidate their nature of synthesis. The potential protein product of the 3 kb RNA will be identified by using antisera against peptides decoded from the nucleotide sequence. The role of this src- related protein in muscle development and function will be studied.