Cathepsin g Degrades Both Glycosylated and Unglycosylated Regions of Lubricin, a Synovial Mucin

Sci Rep. 2020 Mar 6;10(1):4215. doi: 10.1038/s41598-020-61161-5.

Shan Huang ^# ¹, Kristina A Thomsson ^# ¹, Chunsheng Jin ¹ ², Sally Alweddi ¹, André Struglics ³, Ola Rolfson ⁴, Lena I Björkman ⁵, Sebastian Kalamajski ⁶, Tannin A Schmidt ⁷, Gregory D Jay ⁸, Roman Krawetz ⁹ ¹⁰, Niclas G Karlsson ¹¹, Thomas Eisler ¹²

Affiliations

1 Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
2 Science for Life Laboratory, Department of Oncology-Pathology, Clinical Proteomics Mass Spectrometry, Karolinska Institutet, Solna, Sweden.
3 Department of Clinical Sciences Lund, Orthopaedics, Faculty of Medicine, Lund University, Lund, Sweden.
4 Department of Orthopaedics, Institute of Clinical Sciences, The Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
5 Department of Rheumatology and Inflammation Research, Institute of Medicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
6 Department of Molecular Skeletal Biology, Lund University, Lund, Sweden.
7 Biomedical Engineering Department, University of Connecticut Health Centre, Farmington, CT, USA.
8 Department of Emergency Medicine, Warren Alpert Medical School and Division of Biomedical Engineering, School of Engineering, Brown University, Providence, RI, USA.
9 Cell Biology and Anatomy, Cumming School of Medicine, University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta, T2N4N1, Canada.
10 McCaig institute for Bone and Joint Health, University of Calgary, Calgary, Alberta, T2N4N1, Canada.
11 Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden. niclas.karlsson@medkem.gu.se.
12 Department of Clinical Sciences, Danderyd Hospital, Karolinska Institutet, Stockholm, Sweden.
# Contributed equally.

PMID: 32144329 DOI: 10.1038/s41598-020-61161-5

Abstract

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 base 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 all domains of lubricin, including the Mucin domain. Our results suggest that synovial CG may take part in the degradation of lubricin, which could affect the pathological decrease of the lubrication in degenerative joint disease.

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