Vasudevan C, Freund R, Gorga F R
Department of Chemistry and Biochemistry, Duquesne University, Pittsburgh, Pennsylvania 15282, USA.
Virology. 1997 Jul 7;233(2):392-401. doi: 10.1006/viro.1997.8630.
Middle T (mT), the oncogene of murine polyomavirus, causes transformation of rat fibroblasts by activating a number of signal transducing pathways usually used by polypeptide growth factors and their receptors. Here, we report data regarding the activation of signal transducing pathways involving phospholipase D (PL-D). The hydrolysis of phospholipids by PL-D produces phosphatidic acid (PA), a compound with multiple biological effects. The PA content of cells expressing wild-type mT, introduced via a number of different methods, is approximately 50% higher than their untransformed counterparts. This increase in cellular PA content is associated with an approximately 65% increase in PL-D activity in cells expressing wild-type mT. We have also examined the effects of a number of site-directed mutants of mT, on both cellular PA levels and on PL-D activity. Mutants that do not produce mT (Py808A) or that produce a truncated, nonmembrane bound mT (Py1387T) have PA levels similar to that of control cells. Cells expressing the 322YF mutant of mT (which abolishes interaction of mT with phospholipase C gamma1) show increases in both PA levels and PL-D activity that are similar to those seen with wild-type mT. Expression of mutants that abolish the interaction of mT with either shc or with phosphatidylinositol 3-kinase (250YS and 315YF, respectively) cause an increase in PL-D activity comparable to that seen with wild-type mT. However, the PA content of cells expressing these mutants is not elevated. These results suggest that mT causes activation of cellular PL-D, but this activation alone is not sufficient to cause an increase in cellular PA content. Therefore, wild-type mT must affect another, as yet unknown, step in PA metabolism.
小鼠多瘤病毒的致癌基因中T（mT），通过激活许多通常由多肽生长因子及其受体使用的信号转导途径，导致大鼠成纤维细胞发生转化。在此，我们报告了有关涉及磷脂酶D（PL-D）的信号转导途径激活的数据。PL-D对磷脂的水解产生磷脂酸（PA），这是一种具有多种生物学效应的化合物。通过多种不同方法导入的表达野生型mT的细胞中，PA含量比未转化的对应细胞高出约50%。表达野生型mT的细胞中PA含量的这种增加与PL-D活性增加约65%相关。我们还研究了mT的一些定点突变体对细胞PA水平和PL-D活性的影响。不产生mT的突变体（Py808A）或产生截短的、非膜结合mT的突变体（Py1387T）的PA水平与对照细胞相似。表达mT的322YF突变体（该突变体消除了mT与磷脂酶Cγ1的相互作用）的细胞中，PA水平和PL-D活性的增加与野生型mT相似。消除mT与shc或磷脂酰肌醇3激酶相互作用的突变体（分别为250YS和315YF）的表达导致PL-D活性增加，与野生型mT相当。然而，表达这些突变体的细胞中PA含量并未升高。这些结果表明，mT导致细胞PL-D的激活，但仅这种激活不足以导致细胞PA含量增加。因此，野生型mT必须影响PA代谢中的另一个尚未知的步骤。
