Abstract
We investigated interaction of GM3 with N-acetylglucosamine (GlcNAc) termini of N-linked glycans of epidermal growth factor receptor (EGFR), as the underlying mechanism for inhibitory effect of GM3 on EGFR activation, using ldlD cells transfected with EGFR gene. These cells, defective in UDP-Gal/UDP-GalNAc 4-epimerase, are incapable of synthesizing galactose (Gal)-containing glycans, unless Gal is provided in culture (+Gal). Key observations: (1) Expression of GlcNAc termini was high in −Gal cells, and strongly reduced in +Gal cells. (2) Comparative study of inhibitory effect of exogenously-added GM3 on EGFR activation in +Gal versus −Gal cells indicated that higher level of GlcNAc termini on EGFR is correlated with greater inhibitory effect of GM3. (3) GM3-, but not GM1-, coated beads bound to EGFR in lysate of −Gal cells, which have highly exposed GlcNAc termini. Such binding was inhibited in the presence of EDTA, similarly to other carbohydrate-carbohydrate interactions.
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Abbreviations
- EGF:
-
Epidermal growth factor
- EGFR:
-
Epidermal growth factor receptor
- FBS:
-
Fetal bovine serum
- Gal:
-
Galactose
- GlcNAc:
-
N-acetylglucosamine
- ITS:
-
Insulin/transferrin/selenium
- ldlD/EGFR:
-
ldlD cells transfected with EGFR gene
- SDS–PAGE:
-
Sodium dodecyl sulfate polyacrylamide gel electrophoresis
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Acknowledgments
This work was supported in part by NIH National Cancer Institute grant 2 R01 CA080054. The authors thank Monty Krieger for donation of ldlD14 cells, Roger Davis for donation of plasmid pXER, and Steve Anderson for help in preparing the manuscript and figures.
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Special Issue: In Honor of Dr. Robert K. Yu.
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Guan, F., Handa, K. & Hakomori, Si. Regulation of Epidermal Growth Factor Receptor Through Interaction of Ganglioside GM3 with GlcNAc of N-Linked Glycan of the Receptor: Demonstration in ldlD Cells. Neurochem Res 36, 1645–1653 (2011). https://doi.org/10.1007/s11064-010-0379-9
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DOI: https://doi.org/10.1007/s11064-010-0379-9