The elevation of cellular phosphatidic acid levels caused by polyomavirus transformation can be disassociated from the activation of phospholipase D.
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.
Vasudevan
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C
Department of Chemistry and Biochemistry, Duquesne University, Pittsburgh, Pennsylvania 15282, USA.
Freund
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Gorga
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FR
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002472:Cell Transformation, Viral; D004075:Diglycerides / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism* / Q000821:virology; D051379:Mice; D010712:Phosphatidic Acids / Q000378:metabolism*; D010739:Phospholipase D / Q000378:metabolism*; D011120:Polyomavirus / Q000235:genetics / Q000502:physiology*; D051381:Rats
D000952:Antigens, Polyomavirus Transforming; D004075:Diglycerides; D010712:Phosphatidic Acids; D010739:Phospholipase D