N-linked glycosylation regulates human proteinase-activated receptor-1 cell surface expression and disarming via neutrophil proteinases and thermolysin

Xiao, Yu Pei; Morice, Alyn H.; Compton, Steven J.; Sadofsky, Laura

Abstract

Proteinase-activated receptor 1 (PAR1) induces activation of platelet and vascular cells after proteolytic cleavage of its extracellular N terminus by thrombin. In pathological situations, other proteinases may be generated in the circulation and might modify the responses of PAR1 by cleaving extracellular domains. In this study, epitope-tagged wild-type human PAR1 (hPAR1) and a panel of N-linked glycosylation-deficient mutant receptors were permanently expressed in epithelial cells (Kirsten murine sarcoma virus-transformed rat kidney cells and CHO cells). We have analyzed the role of N-linked glycosylation in regulating proteinase activation/disarming and cell global expression of hPAR1. We reported for the first time that glycosylation in the N terminus of hPAR1 downstream of the tethered ligand (especially Asn75) governs receptor disarming to trypsin, thermolysin, and the neutrophil proteinases elastase and proteinase 3 but not cathepsin G. In addition, hPAR1 is heavily N-linked glycosylated and sialylated in epithelial cell lines, and glycosylation occurs at all five consensus sites, namely, Asn35, Asn62, Asn75, Asn250, and Asn259. Removing these N-linked glycosylation sequons affected hPAR1 cell surface expression to varying degrees, and N-linked glycosylation at extracellular loop 2 (especially Asn250) of hPAR1 is essential for optimal receptor cell surface expression and receptor stability.

Citation

Xiao, Y. P., Morice, A. H., Compton, S. J., & Sadofsky, L. (2011). N-linked glycosylation regulates human proteinase-activated receptor-1 cell surface expression and disarming via neutrophil proteinases and thermolysin. Journal of Biological Chemistry, 286(26), 22991-23002. https://doi.org/10.1074/jbc.M110.204271

• Journal Article Type: Article
• Online Publication Date: May 6, 2011
• Publication Date: Jul 1, 2011
• Deposit Date: Nov 13, 2014
• Publicly Available Date: Nov 23, 2017
• Journal: Journal of biological chemistry
• Print ISSN: 0021-9258
• Electronic ISSN: 1083-351X
• Publisher: American Society for Biochemistry and Molecular Biology
• Peer Reviewed: Peer Reviewed
• Volume: 286
• Issue: 26
• Pages: 22991-23002
• DOI: https://doi.org/10.1074/jbc.M110.204271
• Keywords: Cell Biology; Biochemistry; Molecular Biology
• Public URL: https://hull-repository.worktribe.com/output/473014
• Publisher URL: http://www.jbc.org/content/286/26/22991
• Additional Information: Copy of article first published in Journal of biological chemistry, 2011, v.286, issue 26