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N -glycosylation is a common protein post translation modification, which has tremendous structure diversity and wide yet delicate regulation of protein structures and functions. Mass spectrometry-based N -glycoproteomics has become a state-of-the-art pipeline for both qualitative and quantitative characterization of N -glycosylation at the intact N -glycopeptide level, providing comprehensive information of peptide backbones, N -glycosites, monosaccharide compositions, sequence and linkage structures. For high-throughput analysis of large-cohort clinic samples, fast and high-performance separation is indispensable. Here we report our development of 1-h liquid chromatography gradient N -glycoproteomics method and accordingly optimized MS parameters. In the benchmark analysis of cancer and paracancerous tissue of hepatocellular carcinoma, 5,218 intact N -glycopeptides were identified, where 422 site- and structure-specific differential N -glycosylation on 145 N -glycoproteins was observed. The method, representing substantial increase of throughput, can be adopted for fast and efficient analysis of N -glycoproteomes at large scale. [Display omitted] • One-hour N -glycoproteomics with 1-h gradient is developed for fast analysis. • 85 % more IDs was achieved per elution time than previous 4-h gradient. • Fast N -glycoproteomics may find wide application of large-cohort study. [ABSTRACT FROM AUTHOR]

• HA degradation promotes odontoblastic differentiation and cell migration of mouse dental papilla cells. • During odontoblastic differentiation, HYAL2 is highly localized in odontoblasts in vivo. • Hyal2 knockdown leads to the accumulation of HA, which impedes dentinogenesis and cell migration. • Hyal2 silencing significantly affects the cytoskeletal networks and focal adhesion, and inhibits the activation of PI3K/Akt signaling pathway. The coordination between odontoblastic differentiation and directed cell migration of mesenchymal progenitors is necessary for regular dentin formation. The synthesis and degradation of hyaluronan (HA) in the extracellular matrix create a permissive niche that directly regulates cell behaviors. However, the role and mechanisms of HA degradation in dentin formation remain unknown. In this work, we present that HA digestion promotes odontoblastic differentiation and cell migration of mouse dental papilla cells (mDPCs). Hyaluronidase 2 (HYAL2) is responsible for promoting odontoblastic differentiation through degrading HA, while hyaluronidase 1 (HYAL1) exhibits negligible effect. Silencing Hyal2 generates an extracellular environment rich in HA, which attenuates F-actin and filopodium formation and in turn inhibits cell migration of mDPCs. In addition, activating PI3K/Akt signaling significantly rescues the effects of HA accumulation on cytodifferentiation. Taken together, the results confirm the contribution of HYAL2 to HA degradation in dentinogenesis and uncover the mechanism of the HYAL2-mediated HA degradation in regulating the odontoblastic differentiation and migration of mDPCs. [ABSTRACT FROM AUTHOR]