Post-translational processing of bovine chondromodulin-I

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• Chondromodulin-I (ChM-I) is a glycoprotein abundant in fetal cartilage.
• Mature ChM-I is derived from a larger precursor through proteolytic processing.
• Understanding ChM-I processing is crucial for cartilage biology and development.

Purpose of the Study:

• To elucidate the processing pathway and identify the enzymes involved in ChM-I maturation.
• To investigate the intracellular and extracellular events governing ChM-I processing.
• To determine the role of specific cleavage sites and cellular machinery in ChM-I processing.

Main Methods:

• Utilized antipeptide antisera to identify precursor, mature, and processed forms of ChM-I.
• Employed cultured chondrocytes, chondrosarcoma cells, and transfected Chinese hamster ovary (CHO) cells.
• Performed pulse-chase analysis and site-directed mutagenesis of the ChM-I precursor cleavage site.
• Expressed ChM-I in furin-deficient CHO cells to assess furin's role.

Main Results:

• ChM-I precursor is processed intracellularly at the RERR-ELVR site, likely by furin.
• Mature ChM-I is rapidly secreted post-cleavage.
• A smaller, unglycosylated form of ChM-I is generated through an extracellular event, observed specifically in cartilage.
• Mutating the cleavage site or using furin-deficient cells abolished processing.

Conclusions:

• ChM-I processing is a multi-step event involving intracellular furin-mediated cleavage and subsequent extracellular modifications.
• The rapid secretion of mature ChM-I suggests it is not a juxtacrine growth factor.
• The distinct processing pathways highlight tissue-specific modifications of ChM-I.