Transformation by Rous sarcoma virus induces clathrin heavy chain phosphorylation.
We have shown that the heavy chain of clathrin is phosphorylated in chicken embryo fibroblast cells transformed by Rous sarcoma virus, but not in normal cells. Approximately 1 mol of phosphate is bound for every 5 mol of heavy chain in the maximally phosphorylated transformed cells. Two-thirds of the phosphate is on serine and one-third on tyrosine residues. Clathrin heavy chain is a substrate for pp60v-src in vitro. Cleveland analysis of the in vivo and in vitro clathrin heavy chain phosphopeptides, generated by protease V8 digestion, show labeled proteolytic fragments of similar molecular weight, suggesting that pp60v-src could be directly responsible for the in vivo phosphorylation of clathrin. Phosphate is equally incorporated into clathrin in both the unassembled and the assembled clathrin pools, whereas [35S]methionine is preferentially incorporated into the assembled pool. In normal cells, clathrin visualized by immunofluorescent staining appears in a punctate pattern along the membrane surface and concentrated around the nucleus; in transformed cells the perinuclear staining is completely absent. The phosphorylation of clathrin heavy chain in transformed cells may be linked to previously observed transformation-dependent alterations in receptor-mediated endocytosis of ligands such as EGF and thrombin.
Martin-Perez
J
J
Department of Cellular and Developmental Biology, Harvard University, Cambridge, Massachusetts 02138.
Bar-Zvi
D
D
Branton
D
D
Erikson
R L
RL
D000818:Animals; D001358:Avian Sarcoma Viruses / Q000502:physiology*; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral; D002478:Cells, Cultured; D002642:Chick Embryo; D002966:Clathrin / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D005455:Fluorescent Antibody Technique; D010766:Phosphorylation; D011233:Precipitin Tests; D012694:Serine / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D002966:Clathrin; D014443:Tyrosine; D012694:Serine