Array-based functional screening of heparin glycans.
Array methodologies have become powerful tools for interrogation of glycan-protein interactions but have critically lacked the ability to generate cell response data. Here, we report the development of a slide-based array method exemplified by measurement of activation of fibroblast growth factor signaling by heparin saccharides. Heparan sulfate-deficient Swiss 3T3 cells were overlaid onto an aminosilane-coated slide surface onto which heparin saccharides had been spotted and immobilized. The cells were transiently stimulated with FGF2 and immunofluorescence measured to assess downstream ERK1/2 phosphorylation. Activation of this signaling pathway response was restricted to cells exposed to heparin saccharides competent to activate FGF2 signaling. Differential activation of the overlaid cells by different-sized heparin saccharides was demonstrated by quantitative measurement of fluorescence intensity. This "glycobioarray" platform has significant potential as a generic tool for functional glycomics screening.
Centre for Glycobiology, Department of Biochemistry and Cell Biology, Institute of Integrative Biology, The University of Liverpool, Liverpool L69 7ZB, UK.
Tania M
TM
Puvirajesinghe
Yassir A
YA
Ahmed
Andrew K
AK
Powell
David G
DG
Fernig
Scott E
SE
Guimond
Jeremy E
JE
Turnbull
D016475:3T3 Cells; D000818:Animals; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D005346:Fibroblast Growth Factors / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism*; D051379:Mice; D046228:Microarray Analysis / Q000379:methods*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation; D015398:Signal Transduction
D016222:Fibroblast Growth Factor 2; D005346:Fibroblast Growth Factors; D006493:Heparin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3