Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
8452258 | Experimental Cell Research | 2017 | 8 Pages |
Abstract
The highly glycosylated bone sialoprotein (BSP) is an abundant non-collagenous phosphoprotein in bone which enhances osteoblast differentiation and new bone deposition in vitro and in vivo. However, the structural details of its different glycosylation linkages have not been well studied and their functions in bone homeostasis are not clear. Previous studies suggested that the O-glycans, but not the N-glycans on BSP, are highly sialylated. Herein, we employed tandem mass spectrometry (MS/MS) to demonstrate that the N-glycanson the recombinant human integrin binding sialoprotein (rhiBSP) are also enriched in sialic acids (SAs) at their termini. We also identified multiple novel sites of N-glycan modification. Treatment of rhiBSP enhances osteoblast differentiation and mineralization of MC3T3-E1 cells and this effect could be partially reversed by efficient enzymatic removal of its N-glycans. Removal of all terminal SAs has a greater effect in reversing the effect of rhiBSP on osteogenesis, especially on mineralization, suggesting that sialylation at the termini of both N-glycans and O-glycans plays an important role in this regulation. Moreover, BSP-conjugated SAs may affect mineralization via ERK activation of VDR expression. Collectively, our results identified novel N-glycans enriched in SAs on the rhiBSP and demonstrated that SAs at both N- and O-glycans are important for BSP regulation of osteoblast differentiation and mineralization in vitro.
Keywords
ECMOPNBSPOPGHCDDSPPPTMsDMPETDERKNeu5AcMEPERANKLRUNX2high-energy collision dissociationNeu5GcqPCRJnkHPAECc-Jun N-terminal kinaseMAPKPulsed amperometric detectorALPAlkaline phosphataseAktOsteoprotegerinOsteopontinOsteocalcinN-glycolylneuraminic acidN-acetyl neuraminic acidElectron transfer dissociationpost-translational modificationsRankdentin sialophosphoproteinbone sialoproteinRunt-related transcription factor 2High performance anion-exchange chromatographymatrix extracellular phosphoglycoproteinExtracellular matrixHydroxyapatitequantitative real-time PCRPADdentin matrix protein-1protein kinase Bmitogen-activated protein kinaseHPLChigh-performance liquid chromatographyextracellular signal-regulated kinase
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Authors
Lan Xu, Zhenqing Zhang, Xue Sun, Jingjing Wang, Wei Xu, Lv Shi, Jiaojiao Lu, Juan Tang, Jingjing Liu, Xiong Su,