Identification of the motif in versican G3 domain that plays a dominant-negative effect on astrocytoma cell proliferation through inhibiting versicansecretion and binding

This study was designed to investigate the mechanisms by which mutant versi can constructs play a dominant-negative effect on astrocytoma cell prolifer ation. Although a mini-versican or a versican G3 construct promoted growth of U87 astrocytoma cells, a mini-versican lacking epidermal growth factor ( EGF) motifs (versican Delta EGF) and a G3 mutant (G3 Delta EGF) exerted a d ominant-negative effect on cell proliferation. G3 Delta EGF-transfected cel ls formed smaller colonies, arrested cell cycle at G(1) phase, inhibited ex pression of cell cycle proteins cdk4 and cyclin D1, and contained multiple nucleoli, In cell surface binding assays, G3 products expressed in COS-7 ce lls and bacteria bound to U87 cell surface. G3 Delta EGF products exhibited decreased binding activity, but higher levels of G3 Delta EGF products wer e able to inhibit the binding of G3 to the cell surface, G3 Delta EGF expre ssion inhibited secretion of endogenous versican in astrocytoma cells and a lso inhibited the secretion of mini-versican in COS-7 cells co-transfected with the mini-versican and G3 Delta EGF constructs. The effect seems to dep end on the expression efficiency of G3 Delta EGF, and it occurred via the c arbohydrate recognition domain.