Interaction of folate receptor with signaling molecules lyn and G(alpha)(i-3) in detergent-resistant complexes from the ovary carcinoma cell line IGROV1.
Using as a model the ovary carcinoma cell line IGROV1, we analyzed the partitioning of the glycosyl-phosphatidylinositol-anchored folate receptor into lipid rafts based on its relative detergent insolubility, with a focus on physically and functionally associated signaling molecules. A variable amount (40-60%) of folate receptor was found in low-density Triton X-100 insoluble complexes together with subunits of heterotrimeric G-proteins and the src-family non-receptor tyrosine kinases p53-56 lyn. In the same fraction the structural component of caveolae, caveolin, was not detected at the protein level, although the corresponding mRNA was detected in trace amounts. Comodulation of folate receptor and signalling molecules was observed in the detergent-insoluble complexes during cell proliferation or induced by phosphatidylinositol-specific phospholipase C treatment or by interaction with anti-folate receptor monoclonal antibodies. Moreover, complexes of folate receptor, lyn and the G(&agr;)(i-3) subunit were immunoprecipitated using either anti-folate receptor or anti-lyn antibodies. In vitro kinase assay of the immunoprecipitates revealed stimulation of phosphorylation of common and specific proteins. In particular, the p53 form of lyn appeared to be enriched and phosphorylated in the anti-folate receptor MOv19 monoclonal antibody immunoprecipitate, whereas a 40 kDa band common to anti-folate receptor and anti-lyn immunoprecipitates was the phosphorylated form of the G(&agr;)(i-3) subunit. These findings point to the functional interaction between folate receptor and associated signaling molecules.
Miotti
S
S
Unit of Molecular Therapies, Department of Experimental Oncology, Istituto Nazionale per lo Studio e la Cura dei Tumori, Milan, Italy. miotti@istitutotumori.mi.it.
Bagnoli
M
M
Tomassetti
A
A
Colnaghi
M I
MI
Canevari
S
S
D002352:Carrier Proteins / Q000378:metabolism*; D051242:Caveolin 1; D022461:Caveolins; D003902:Detergents; D005260:Female; D058973:Folate Receptors, GPI-Anchored; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D017830:Octoxynol; D010051:Ovarian Neoplasms / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011956:Receptors, Cell Surface; D015398:Signal Transduction; D012995:Solubility; D014407:Tumor Cells, Cultured; D019061:src-Family Kinases / Q000378:metabolism*
C495540:CAV1 protein, human; D002352:Carrier Proteins; D051242:Caveolin 1; D022461:Caveolins; D003902:Detergents; D058973:Folate Receptors, GPI-Anchored; D008565:Membrane Proteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D017830:Octoxynol; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go