猫c-fes原癌基因的分子克隆及嵌合转化基因的构建。

Molecular cloning of the feline c-fes proto-oncogene and construction of a chimeric transforming gene.

作者信息

Verbeek J S, van den Ouweland A M, Schalken J A, Roebroek A J, Onnekink C, Bloemers H P, van de Ven W J

出版信息

Gene. 1985;35(1-2):33-43. doi: 10.1016/0378-1119(85)90155-6.

Abstract

The feline c-fes proto-oncogene, different parts of which were captured in feline leukemia virus (FeLV) to generate the transforming genes (v-fes) of the Gardner-Arnstein (GA) strain of feline sarcoma virus (FeSV) and the Snyder-Theilen strain (ST) of FeSV, was cloned and its genetic organization determined. Southern blot analysis revealed that the c-fes genetic sequences were distributed discontinuously and colinearly with the v-fes transforming gene over a DNA region of around 12.0 kb. Using cloned c-fes sequences, complementation of GA-FeSV transforming activity was studied. Upon replacement of the 3' half of v-fesGA with homologous feline c-fes sequences and transfection of the chimeric gene, morphological transformation was observed. Immunoprecipitation analysis of these transformed cells revealed expression of high Mr fusion proteins. Phosphorylation of these proteins was observed in an in vitro protein kinase assay, and tyrosine residues appeared to be involved as acceptor amino acid.

摘要

猫c-fes原癌基因被克隆并确定了其遗传结构，该基因的不同部分被猫白血病病毒（FeLV）捕获，从而产生了加德纳-阿恩斯坦（GA）株猫肉瘤病毒（FeSV）和斯奈德-泰伦（ST）株FeSV的转化基因（v-fes）。Southern印迹分析表明，c-fes基因序列在约12.0 kb的DNA区域内与v-fes转化基因不连续且共线分布。利用克隆的c-fes序列，研究了GA-FeSV转化活性的互补作用。用同源猫c-fes序列取代v-fesGA的3'端一半并转染嵌合基因后，观察到形态转化。对这些转化细胞的免疫沉淀分析显示出高分子量融合蛋白的表达。在体外蛋白激酶试验中观察到这些蛋白的磷酸化，酪氨酸残基似乎作为受体氨基酸参与其中。