FIG 3.

AlHV-sema is a 93-kDa secreted glycoprotein. (A) Immunoblot analysis of sema7A and AlHV-sema. HeLa cells were transfected with pEX-sema7A, pTA3Fc, pTA3V5, or empty vector. At 24 h, the proteins in the supernatant were analyzed by immunoblotting with anti-human IgG1 goat polyserum or anti-V5 MAb. Supernatants were left untreated (−) or subjected to deglycosylation treatment (PNGase F, O-glycosidase, neuraminidase, β1-4 galactosidase, β-N-acetylglucosaminidase) before SDS-PAGE and immunoblotting. The positions of molecular size markers are indicated in kilodaltons. (B) Transient expression of sema7A or AlHV-sema was performed in HeLa cells in the presence or absence of tunicamycin (Tunic.) or brefeldin A (Bref.A), and both secreted (Supernatant) or cell-associated (Cells) proteins were analyzed by immunoblotting.