title: Polyamines are essential for cell transformation by pp60v-src: delineation of molecular events relevant for the transformed phenotype.
abstract: Ornithine decarboxylase (ODC), the key enzyme of polyamine biosynthesis, becomes upregulated during cell proliferation and transformation. Here we show that intact ODC activity is needed for the acquisition of a transformed phenotype in rat 2R cells infected with a temperature-sensitive mutant of Rous sarcoma virus. Addition of the ODC inhibitor alpha-difluoromethyl ornithine (DFMO) to the cells (in polyamine-free medium) before shift to permissive temperature prevented the depolymerization of filamentous actin and morphological transformation. Polyamine supplementation restored the transforming potential of pp60v-src. DFMO did not interfere with the expression of pp60v-src or its in vitro tyrosine kinase activity. The tyrosine phosphorylation of most cellular proteins, including ras GAP, did not either display clear temperature- or DFMO-sensitive changes. A marked increase was, however, observed in the tyrosine phosphorylation of phosphatidylinositol 3-kinase and proteins of 33 and 36 kD upon the temperature shift, and these hyperphosphorylations were partially inhibited by DFMO. A DFMO-sensitive increase was also found in the total phosphorylation of calpactins I and II. The well-documented association of GAP with the phosphotyrosine-containing proteins p190 and p62 did not correlate with transformation, but a novel 42-kD tyrosine phosphorylated protein was complexed with GAP in a polyamine- and transformation-dependent manner. Further, tyrosine phosphorylated proteins of 130, 80/85, and 36 kD were found to coimmunoprecipitate with pp60v-src in a transformation-related manner. Altogether, this model offers a tool for sorting out the protein phosphorylations and associations critical for the transformed phenotype triggered by pp60v-src, and implicates a pivotal role for polyamines in cell transformation.
authors.affiliation: Department of Pathology, University of Helsinki, Finland.
authors.forename: E
authors.initials: E
authors.lastname: Hölttä
authors.forename: M
authors.initials: M
authors.lastname: Auvinen
authors.forename: L C
authors.initials: LC
authors.lastname: Andersson
mesh_terms: D008841:Actin Cytoskeleton / Q000648:ultrastructure; D000199:Actins / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002472:Cell Transformation, Viral; D000518:Eflornithine / Q000494:pharmacology; D020690:GTPase-Activating Proteins; D015688:Oncogene Protein pp60(v-src) / Q000502:physiology*; D009955:Ornithine Decarboxylase / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D019000:Phosphotyrosine; D011073:Polyamines / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism; D020703:ras GTPase-Activating Proteins
chemical_list: D000199:Actins; D020690:GTPase-Activating Proteins; D010750:Phosphoproteins; D011073:Polyamines; D011506:Proteins; D020703:ras GTPase-Activating Proteins; D019000:Phosphotyrosine; D014443:Tyrosine; D010770:Phosphotransferases; D019869:Phosphatidylinositol 3-Kinases; D015688:Oncogene Protein pp60(v-src); D009955:Ornithine Decarboxylase; D000518:Eflornithine