Roles of aggrecan domains in biosynthesis, modification by glycosaminoglycans and product secretion

Aggrecan is a member of the chondroitin sulphate (CS) proteoglycan family, which also includes versican/PG-M, neurocan and brevican. Members of this f amily exhibit structural similarity: a G1 domain at the N-terminus and a G3 domain at the C-terminus, with a central sequence for modification by CS c hains. A unique feature of aggrecan is the insertion of three additional do mains. an inter-globular domain (IGD), a G2 domain and a keratan sulphate ( KS) domain (sequence modified by KS chains), between the G1 domain and the CS domain (sequence modified by CS chains). The G1 and G3 domains have been implicated in product secretion, but G2, although structurally similar to the tandem repeats of G1, performs an unknown function. To define the funct ions of each aggrecan domain in product processing, we cloned and expressed these domains in various combinations in COS-7 cells. The results indicate d that the G3 domain enhanced product secretion, alone or in combination wi th the KS or CS domain: and promoted glycosaminoglycan (GAG) chain attachme nt. Constructs containing the G1 domain were not secreted. Addition of a CS domain sequence to G1 reduced this inhibition, but GAG chain attachment wa s still decreased. The potential GAG chain attachment site in the IGD was o ccupied by GAGs, and IGD product was secreted efficiently. The KS domain wa s modified by GAG chains and secreted. Finally, the G2 domain was expressed but not secreted, and inhibited secretion of the IGD when expressed as an IGD-G2 combination.