Intracellular retention of membrane-anchored v-sis protein abrogates autocrine signal transduction.
An important question regarding autocrine transformation by v-sis is whether intracellularly activated PDGF receptors are sufficient to transform cells or whether activated receptor-ligand complexes are required at the cell surface. We have addressed this question by inhibiting cell surface transport of a membrane-anchored v-sis protein utilizing the ER retention signal of the adenoviral transmembrane protein E3/19K. A v-sis fusion protein containing this signal was retained within the cell and not transported to the cell surface as confirmed by immunofluorescent localization experiments. Also, proteolytic maturation of this protein was suppressed, indicating inefficient transport to post-Golgi compartments of the secretory pathway. When compared with v-sis proteins lacking a functional retention signal, the ER-retained protein showed a diminished ability to transform NIH 3T3 cells, as measured by the number and size of foci formed. In newly established cell lines, the ER-retained protein did not down-regulate PDGF receptors. However, continued passage of these cells selected for a fully transformed phenotype exhibiting downregulated PDGF receptors and proteolytically processed v-sis protein. These results indicate that productive autocrine interactions occur in a post-ER compartment of the secretory pathway. Transport of v-sis protein beyond the Golgi correlated with acquisition of the transformed phenotype. Furthermore, suramin treatment reversed transformation and upregulated the expression of cell surface PDGF receptors, suggesting an important role for receptor-ligand complexes localized to the cell surface.
Lee
B A
BA
Department of Biology, University of California, San Diego, La Jolla 92093-0322.
Donoghue
D J
DJ
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002462:Cell Membrane / Q000378:metabolism*; D002471:Cell Transformation, Neoplastic / Q000187:drug effects; D015536:Down-Regulation; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D020575:Oncogene Proteins v-sis; D011956:Receptors, Cell Surface / Q000378:metabolism; D017479:Receptors, Platelet-Derived Growth Factor; D015682:Retroviridae Proteins, Oncogenic / Q000378:metabolism*; D012842:Sarcoma Virus, Woolly Monkey; D015398:Signal Transduction; D013498:Suramin / Q000494:pharmacology; D015854:Up-Regulation
D020575:Oncogene Proteins v-sis; D011956:Receptors, Cell Surface; D015682:Retroviridae Proteins, Oncogenic; D013498:Suramin; D017479:Receptors, Platelet-Derived Growth Factor