Intracellular targeting of pp60src expression: localization of v-src to adhesion plaques is sufficient to transform chicken embryo fibroblasts.
To define the effects of pp60v-src activity at different intracellular sites, we have constructed chimeric molecules which target the pp60v-src kinase to specific intracellular locations. pp60v-src was targeted to the nucleus by insertion of the SV40 large T antigen nuclear localization signal. Nuclear pp60v-src was active as a tyrosine kinase and phosphorylated nuclear proteins at tyrosine. However, cells expressing the nuclear pp60v-src were phenotypically normal by a number of criteria, and nuclear src kinase did not induce the expression of an mRNA (CEF-4) whose induction is characteristic of transformation by wild-type v-src. pp60v-src was targeted to perinuclear membranes by fusion to rat growth hormone and vesicular stomatitis G protein sequences. Cells expressing this chimeric molecule were phenotypically normal by most criteria. However the perinuclear src protein did induce elevated levels of CEF-4 mRNA, indicating that the v-src kinase expressed at this site induces partial transformation. The v-src and activated c-src kinases were targeted to adhesion plaques by fusion to the talin-binding sequence of vinculin. Cells expressing these fusion proteins were transformed by morphological, physiological and biochemical criteria, although the foci induced by these viruses were distinct from those induced by wild-type v-src. A chimeric protein which targeted c-src to adhesion plaques was not transforming. Thus targeting pp60src to adhesion plaques, although not sufficient to activate the transforming capacity of c-src, is sufficient to allow transformation by v-src.
Liebl
E C
EC
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Martin
G S
GS
D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000032:analysis / Q000235:genetics; D001692:Biological Transport; D002448:Cell Adhesion; D002455:Cell Division; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D002642:Chick Embryo; D003847:Deoxyglucose / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D016391:Genes, src; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D015688:Oncogene Protein pp60(v-src) / Q000235:genetics* / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013539:Simian virus 40 / Q000235:genetics; D014162:Transfection
D000952:Antigens, Polyomavirus Transforming; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D003847:Deoxyglucose; D011505:Protein-Tyrosine Kinases; D015688:Oncogene Protein pp60(v-src)