Ganglioside GD3 enhances adhesion signals and augments malignant properties of melanoma cells by recruiting integrins to glycolipid-enriched microdomains.
Ganglioside GD3 is widely expressed in human malignant melanoma cell lines and tumors. Previously, we reported that GD3+ cells show stronger tyrosine phosphorylation of focal adhesion kinase (FAK), p130(Cas), and paxillin when treated with fetal calf serum than GD3- cells. In this study, we analyzed the changes in the signals mediated by the interaction between integrins and extracellular matrices (ECM) to clarify how GD3 enhances cell signals in the vicinity of the cell membrane. An adhesion assay with a real time cell electronic sensing system revealed that GD3+ cells had stronger adhesion to all extracellular matrices examined. In particular, GD3+ cells attached more strongly to collagen type I and type IV than controls. Correspondingly, they showed stronger tyrosine phosphorylation of FAK and paxillin during adhesion to collagen type I. In the floating pattern of detergent extracts, a high level of integrin beta1 was found in glycolipid-enriched microdomain (GEM)/rafts in GD3+ cells before adhesion, whereas a smaller amount of integrin beta1 was detected in the GEM/rafts of controls. Some phosphorylated forms of FAK as well as total FAK were found in GEM/rafts during cell adhesion only in GD3+ cells. Another signal consisting of integrin-linked kinase/Akt was also activated during adhesion more strongly in GD3+ cells than in controls. In double stained GD3+ cells, GD3 and integrin beta1 co-localized at the focal adhesion with a punctate pattern. All these results suggested that integrins assembled and formed a cluster in GEM/rafts, leading to the enhanced signaling and malignant properties under GD3 expression.
Ohkawa
Yuki
Y
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065.
Miyazaki
Sayaka
S
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065.
Hamamura
Kazunori
K
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065.
Kambe
Mariko
M
Department of Biomedical Sciences, College of Life and Health Sciences, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan; Health Science Hills, College of Life and Health Sciences, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan.
Miyata
Maiko
M
Department of Biomedical Sciences, College of Life and Health Sciences, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan.
Tajima
Orie
O
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065; Department of Biomedical Sciences, College of Life and Health Sciences, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan.
Ohmi
Yuhsuke
Y
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065.
Yamauchi
Yoshio
Y
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065.
Furukawa
Koichi
K
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065. Electronic address: koichi@med.nagoya-u.ac.jp.
Furukawa
Keiko
K
Department of Biochemistry II, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-0065; Department of Biomedical Sciences, College of Life and Health Sciences, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan; Health Science Hills, College of Life and Health Sciences, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan. Electronic address: keikofu@isc.chubu.ac.jp.
D002448:Cell Adhesion / Q000235:genetics; D045744:Cell Line, Tumor; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D024141:Collagen Type IV / Q000235:genetics / Q000378:metabolism; D050719:Crk-Associated Substrate Protein / Q000235:genetics / Q000378:metabolism; D005109:Extracellular Matrix / Q000235:genetics / Q000378:metabolism; D051417:Focal Adhesion Kinase 1 / Q000235:genetics / Q000378:metabolism; D022001:Focal Adhesions / Q000235:genetics / Q000378:metabolism*; D005732:Gangliosides / Q000096:biosynthesis* / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006017:Glycolipids / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D019012:Integrin beta1 / Q000235:genetics / Q000378:metabolism*; D008545:Melanoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D021962:Membrane Microdomains / Q000235:genetics / Q000378:metabolism*; D051419:Paxillin / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000235:genetics; D015398:Signal Transduction
D024042:Collagen Type I; D024141:Collagen Type IV; D050719:Crk-Associated Substrate Protein; D005732:Gangliosides; D006017:Glycolipids; D019012:Integrin beta1; C496061:PXN protein, human; D051419:Paxillin; C026226:ganglioside, GD3; D051417:Focal Adhesion Kinase 1; C496050:PTK2 protein, human