Zavala Guillermo, Pretto Carla, Chow Yen-Hung J, Jones LeeAnn, Alberti Alberto, Grego Elena, De las Heras Marcelo, Palmarini Massimo
Department of Medical Microbiology and Parasitology, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.
Virology. 2003 Jul 20;312(1):95-105. doi: 10.1016/s0042-6822(03)00205-8.
Expression of the JSRV envelope (Env) is sufficient to transform immortalized rodent fibroblasts. A putative docking site for the PI3-K kinase (Y(590)-X-X-M(593)) in the cytoplasmic tail of the transmembrane domain of the JSRV Env is a major determinant of viral-induced cell transformation. Akt is constitutively phosphorylated in rodent fibroblasts transformed by the JSRV Env. However, recent data suggest that Y590 and M593 are not necessary for JSRV Env-induced transformation of the immortalized chicken fibroblasts cell line DF-1. In this study we found that JSRV-induced transformation of DF-1 cells is Akt-independent. In addition, a replication-competent avian vector expressing the JSRV Env (RCASBP(A)+JE) was also able to induce transformation of primary chicken embryo fibroblasts (CEF). Vectors expressing JSRV Env Y590 mutants were still able to induce CEF cells transformation but not as efficiently as the vectors expressing the wild-type Env. In CEF cells, as in DF-1 cells, only the expression of the wild-type Env induced constitutive phosphorylation of Akt. Thus, in chicken cells, the degree of transformation induced by the JSRV Env is maximum in the presence of Y590 and Akt phosphorylation. We addressed the significance of Akt phosphorylation in rat 208F cells transformed by the JSRV Env and showed that Akt is indeed activated and shows kinase activity. Inhibitors of the PI-3K/Akt pathway reproducibly decreased the transformation efficiency of the JSRV Env. In vivo, we found phosphorylated Akt only in nasal tumors induced by the enzootic nasal tumor virus (ENTV), a JSRV-related beta-retrovirus. No evidence of Akt phosphorylation was found in lung tumor sections of sheep affected by pulmonary adenocarcinoma. As a whole, these results suggest that the activation of the PI-3K/Akt pathway contributes to the process of JSRV-induced cell transformation but most likely is not the primary determinant both in vitro and in vivo.
禽肉瘤病毒囊膜（Env）的表达足以使永生化啮齿动物成纤维细胞发生转化。禽肉瘤病毒Env跨膜结构域胞质尾中一个假定的磷脂酰肌醇-3激酶（PI3-K激酶）对接位点（Y(590)-X-X-M(593)）是病毒诱导细胞转化的主要决定因素。在由禽肉瘤病毒Env转化的啮齿动物成纤维细胞中，Akt持续磷酸化。然而，最近的数据表明，Y590和M593对于禽肉瘤病毒Env诱导永生化鸡成纤维细胞系DF-1的转化并非必需。在本研究中，我们发现禽肉瘤病毒诱导的DF-1细胞转化不依赖Akt。此外，表达禽肉瘤病毒Env的复制型禽载体（RCASBP(A)+JE）也能够诱导原代鸡胚成纤维细胞（CEF）发生转化。表达禽肉瘤病毒Env Y590突变体的载体仍能够诱导CEF细胞转化，但效率不如表达野生型Env的载体。在CEF细胞中，与DF-1细胞一样，只有野生型Env的表达诱导Akt的持续磷酸化。因此，在鸡细胞中，在存在Y590和Akt磷酸化的情况下，禽肉瘤病毒Env诱导的转化程度最大。我们探讨了Akt磷酸化在由禽肉瘤病毒Env转化的大鼠208F细胞中的意义，并表明Akt确实被激活并具有激酶活性。PI-3K/Akt途径的抑制剂可重复性地降低禽肉瘤病毒Env的转化效率。在体内，我们仅在由地方流行性鼻肿瘤病毒（ENTV，一种与禽肉瘤病毒相关的β逆转录病毒）诱导的鼻肿瘤中发现磷酸化的Akt。在受肺腺癌影响的绵羊肺肿瘤切片中未发现Akt磷酸化的证据。总体而言，这些结果表明PI-3K/Akt途径的激活有助于禽肉瘤病毒诱导的细胞转化过程，但很可能在体外和体内都不是主要决定因素。
