Jong S M, Wang L H
Department of Microbiology, Mount Sinai School of Medicine, New York, New York 10029-6574.
J Virol. 1990 Dec;64(12):5997-6009. doi: 10.1128/JVI.64.12.5997-6009.1990.
The transforming protein P68gag-ros of avian sarcoma virus UR2 is a transmembrane tyrosine protein kinase molecule with the gag portion protruding extracellularly. To investigate the role of the gag moiety in the biochemical properties and biological functions of the P68gag-ros fusion protein, retroviruses containing the ros coding sequence of UR2 were constructed and analyzed. The gag-free ros protein was expressed from one of the mutant retroviruses at a level 10 to 50% of that of the wild-type UR2. However, the gag-free ros-containing viruses were not able to either transform chicken embryo fibroblasts or induce tumors in chickens. The specific tyrosine protein kinase activity of gag-free ros protein is about 10- to 20-fold reduced as judged by in vitro autophosphorylation. The gag-free ros protein is still capable of associating with membrane fractions including the plasma membrane, indicating that sequences essential for recognition and binding membranes must be located within ros. Upon passages of the gag-free mutants, transforming and tumorigenic variants occasionally emerged. The variants were found to have regained the gag sequence fused to the 5' end of the ros, apparently via recombination with the helper virus or through intramolecular recombination between ros and upstream gag sequences in the same virus construct. All three variants analyzed code for gag-ros fusion protein larger than 68 kDa. The gag-ros recombination junction of one of the transforming variants was sequenced and found to consist of a p19-p10-p27-ros fusion sequence. We conclude that the gag sequence is essential for the transforming activity of P68gag-ros but is not important for its membrane association.
禽肉瘤病毒UR2的转化蛋白P68gag-ros是一种跨膜酪氨酸蛋白激酶分子，其gag部分突出于细胞外。为了研究gag部分在P68gag-ros融合蛋白的生化特性和生物学功能中的作用，构建并分析了含有UR2的ros编码序列的逆转录病毒。无gag的ros蛋白在一种突变逆转录病毒中表达，其水平为野生型UR2的10%至50%。然而，不含gag的含ros病毒既不能转化鸡胚成纤维细胞，也不能在鸡中诱导肿瘤。通过体外自磷酸化判断，无gag的ros蛋白的特异性酪氨酸蛋白激酶活性降低了约10至20倍。无gag的ros蛋白仍能与包括质膜在内的膜组分结合，这表明识别和结合膜所必需的序列一定位于ros内。在无gag的突变体传代后，偶尔会出现转化和致瘤变体。发现这些变体重新获得了与ros的5'端融合的gag序列，显然是通过与辅助病毒重组或通过同一病毒构建体中ros与上游gag序列之间的分子内重组。分析的所有三个变体编码的gag-ros融合蛋白都大于68 kDa。对其中一个转化变体的gag-ros重组连接点进行了测序，发现其由p19-p10-p27-ros融合序列组成。我们得出结论，gag序列对于P68gag-ros的转化活性是必不可少的，但对于其膜结合并不重要。
