Sialidase treatment exposes the beta1-integrin active ligand binding site on HL60 cells and increases binding to fibronectin.
The migration of neutrophils from the circulation to areas of inflammation is the result of the sequential activation of multiple cellular adhesion molecules. beta1-Integrins are cell surface glycoproteins and the class of adhesion molecules responsible for binding to the extracellular matrix. The goal of this study was to determine the contribution of glycosylation, specifically the presence of sialic acid, to beta1-integrin adhesion in a neutrophil model. beta1-Integrins on differentiated HL60 cells were remodeled by treatment with the exoglycosidases, sialidase and beta-galactosidase. beta1-Integrin activity was determined by measuring adherence to the extracellular matrix protein fibronectin. The expression of beta1-integrins, beta2-integrins and activated beta1-integrins was determined by flow cytometry. Remodeling of beta1-integrins by treatment with sialidase increased adhesion by greater than 1,000%. Flow cytometric analysis of remodeled beta1-integrins demonstrated an increased expression of the activated beta1-integrin, but only minor increases in the expression of total beta1- and beta2-integrins. We postulate that glycosidase treatment increases adhesion and expression of activated beta1-integrins by exposure of the normally hidden ligand-binding site. The glycosylation of beta1-integrins on neutrophils may act to hide the ligand-binding site in unstimulated cells thereby contributing to the affinity modulation observed in neutrophil beta1-integrin function.
Pretzlaff
R K
RK
Division of Critical Care Medicine, Children's Hospital Medical Center, Cincinnati, OH 45229, USA. PretzlaffR@aol.com
Xue
V W
VW
Rowin
M E
ME
D001665:Binding Sites / Q000187:drug effects / Q000502:physiology; D015153:Blotting, Western; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002454:Cell Differentiation; D004121:Dimethyl Sulfoxide; D005109:Extracellular Matrix / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism*; D005434:Flow Cytometry; D018922:HL-60 Cells; D006801:Humans; D019012:Integrin beta1 / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism*; D008024:Ligands; D009439:Neuraminidase / Q000494:pharmacology*; D011233:Precipitin Tests; D012997:Solvents; D001616:beta-Galactosidase / Q000494:pharmacology
D005353:Fibronectins; D019012:Integrin beta1; D008024:Ligands; D012997:Solvents; D009439:Neuraminidase; D001616:beta-Galactosidase; D004121:Dimethyl Sulfoxide