A close association of GM3 with c-Src and Rho in GM3-enriched microdomains at the B16 melanoma cell surface membrane: a preliminary note.
B16 melanoma is characterized by high content of GM3 ganglioside, which has been recognized as a melanoma-associated antigen defined by specific monoclonal antibodies. We report now that GM3 is present predominantly (>90%) in the 1% Triton X-100-insoluble, low-density microvesicular fraction ("detergent-insoluble glycosphingolipid-enriched microdomain"; DIGEM) separated on sucrose density-gradient centrifugation. Associated with DIGEM, many signal transducer molecules such as c-Src, FAK, and the low-molecular-weight G-proteins Rho A and H-Ras were also found. Rho A and FAK were found in part, and PLC-beta2 and G alphas were found exclusively, in the high-density fraction. Immunoprecipitation of GM3 present in DIGEM by anti-GM3 antibody DH2, followed by Western blotting, revealed co-precipitation of Rho A and c-Src with GM3. These findings suggest (i) a specific organization of GM3 in close association with Rho A and c-Src within DIGEM at the melanoma cell surface; and (ii) such organizational units may be directly involved in signal transduction, in which glycosphingolipids receive signals which are subsequently transduced by associated transducer molecules.
Yamamura
S
S
Department of Pathobiology, University of Washington and Pacific Northwest Research Foundation, Seattle 98122, USA.
Handa
K
K
Hakomori
S
S
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D005679:G(M3) Ganglioside / Q000032:analysis*; D019204:GTP-Binding Proteins / Q000032:analysis*; D008546:Melanoma, Experimental / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D014407:Tumor Cells, Cultured; D019061:src-Family Kinases / Q000032:analysis*
D005679:G(M3) Ganglioside; D019061:src-Family Kinases; D019204:GTP-Binding Proteins