Rolfson, O. | bioRxiv Science

bioRxiv2019

Core-2 O-glycans are required for Galectin-3 interaction with the osteoarthritis related protein lubricin

Synovial fluid lubricin (proteoglycan 4) is a mucin-type O-linked glycosylated (60% of the mass) biological lubricant involved in osteoarthritis (OA) development. Lubricin has been reported to be cross-linked by synovial galectin-3 on the lubricating articular surface. Here, we confirm that binding to galectin-3 depended on core-2 O-linked glycans, where surface plasmon resonance of a recombinant lubricin (rhPRG4) devoid of core-2 structures lacked binding capacity to recombinant galectin-3. Both galectin-3 levels and interactions with synovial lubricin were found to be decreased in late-stage OA patients coinciding with an increase of truncated and less sialylated core 1 O-glycans. These data suggest a defect cross-linking of surface active molecules in OA and provides novel insights into OA molecular pathology.

biochemistry↗

bioRxiv2019

Cathepsin G Degrades Synovial Fluid Lubricin: Relevance For Osteoarthritis Pathogenesis

Lubricin (PRG4) is a mucin type protein that plays an important role in maintaining normal joint function by providing lubrication and chondroprotection. Improper lubricin modification and degradation has been observed in idiopathic osteoarthritis (OA), while the detailed mechanism still remains unknown. We hypothesized that the protease cathepsin G (CG) may participate in degrading lubricin in synovial fluid (SF). The presence of endogenous CG in SF was confirmed in 16 patients with knee OA. Recombinant human lubricin (rhPRG4) and native lubricin purified from the SF of patients were incubated with exogenous CG and lubricin degradation was monitored using western blot, staining by Coomassie or Periodic Acid-Schiff in gels, and with proteomics. Full length lubricin ([~]300 kDa), was efficiently digested with CG generating a 25-kDa protein fragment, originating from the densely glycosylated mucin domain ([~]250 kDa). The 25-kDa fragment was present in the SF from OA patients, and the amount was increased after incubation with CG. A CG digest of rhPRG4 revealed 135 peptides and 72 glycopeptides, and confirmed that the protease could cleave in different domains of lubricin. Our results suggest that synovial CG may take part in the degradation of lubricin, which could affect the lubrication of OA joints.

pathology↗