Role of gangliosides in the association of ErbB2 with lipid rafts in mammary epithelial HC11 cells.
We analyzed the role of gangliosides in the association of the ErbB2 receptor tyrosine-kinase (RTK) with lipid rafts in mammary epithelial HC11 cells. Scanning confocal microscopy experiments revealed a strict ErbB2-GM3 colocalization in wild-type cells. In addition, analysis of membrane fractions obtained using a linear sucrose gradient showed that ErbB2, epidermal growth factor receptor (EGFR) and Shc-p66 (proteins correlated with the ErbB2 signal transduction pathway) were preferentially enriched in lipid rafts together with gangliosides. Blocking of endogenous ganglioside synthesis by (+/-)-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol hydrochloride ([D]-PDMP) induced a drastic cell-surface redistribution of ErbB2, EGFR and Shc-p66, within the Triton-soluble fractions, as revealed by linear sucrose-gradient analysis. This redistribution was partially reverted when exogenous GM3 was added to ganglioside-depleted HC11 cells. The results point out the key role of ganglioside GM3 in retaining ErbB2 and signal-transduction-correlated proteins in lipid rafts.
Sottocornola
Elena
E
Institute of General Physiology and Biological Chemistry, University of Milan, Italy.
Misasi
Roberta
R
Mattei
Vincenzo
V
Ciarlo
Laura
L
Gradini
Roberto
R
Garofalo
Tina
T
Berra
Bruno
B
Colombo
Irma
I
Sorice
Maurizio
M
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism; D005679:G(M3) Ganglioside / Q000502:physiology*; D015151:Immunoblotting; D047468:Immunoprecipitation; D008321:Mammary Glands, Animal / Q000166:cytology*; D021962:Membrane Microdomains / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D009025:Morpholines / Q000494:pharmacology; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism*; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1
D048868:Adaptor Proteins, Signal Transducing; D002784:Cholesterol; D066246:ErbB Receptors; D005679:G(M3) Ganglioside; D009025:Morpholines; D018719:Erb-b2 Receptor Tyrosine Kinases; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; C528034:Shc1 protein, mouse; C033110:RV 538