Osteopontin O-glycosylation contributes to its phosphorylation and cell-adhesion properties.

Abstract

OPN (osteopontin) is a multiphosphorylated extracellular glycoprotein, which has important roles in bone remodelling, inflammation and cancer metastasis. OPN regulates cell spreading and adhesion primarily through its association with several integrins such as αvβ3, and its phosphorylation affects these processes. However, the mechanism by which OPN O-glycosylation affects these processes is not completely understood. In the present study, we demonstrated that OPN O-glycosylation self-regulates its biological activities and also affects its phosphorylation status. We prepared two recombinant OPNs, WT (wild-type)-OPN and mutant OPN (ΔO-OPN), which lacks five O-glycosylation sites at a threonine/proline-rich region. O-glycan defects in OPN increased its phosphorylation level, as observed by dephosphorylation assays. Moreover, compared with WT-OPN, ΔO-OPN exhibited enhanced cell spreading and adhesion activities and decreased associations with β1 integrins. This suggested that defects in O-glycans in OPN altered these activities, and that β1 integrins have a less important role in adhesion to ΔO-OPN. The cell-adhesion activity of dephosphorylated ΔO-OPN was higher than the cell-adhesion activities of ΔO-OPN and dephosphorylated WT-OPN. This suggested that some of the phosphorylation in ΔO-OPN caused by O-glycan defects and O-glycans of OPN suppressed the OPN cell-adhesion activity. Thus functional activities of OPN can be determined by the combined glycosylation and phosphorylation statuses and not by either status alone.

• Authors: Yoshinobu Kariya, Mayumi Kanno, Kana Matsumoto-Morita, Midori Konno, Yoshiki Yamaguchi, Yasuhiro Hashimoto
• Journal: The Biochemical journal (Biochem J) Vol. 463 Issue 1 Pg. 93-102 (Oct 01 2014) ISSN: 1470-8728 [Electronic] England
• PMID: 25000122 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Integrin alphaVbeta3
• Osteopontin

Topics

• Cell Adhesion (physiology)
• Cell Line, Tumor
• Glycosylation
• HEK293 Cells
• Humans
• Integrin alphaVbeta3 (genetics, metabolism)
• Osteopontin (genetics, metabolism)
• Phosphorylation (physiology)