Fig. 2.

Glycosylation of the Rem C terminus is required for Rem function but not for cleavage. (A) Western blotting of 293T cells transfected with 250 ng of pHMRluc, 250 ng of pGL3, and 5.5 μg of wild-type rem or glycosylation-site mutants as indicated. The three forms of Rem detected with SP-specific antibody (upper arrows) include Rem glycosylated at positions 127 and 143 (lane 2; 38-kDa band), at amino acid 143 or 127 only (lanes 3 and 4), or at neither position (lane 5). (B) Rem function requires at least one functional glycosylation site. Transient transfections of 293T cells were performed using the indicated amount of wild-type rem or glycosylation-site mutant constructs. Luciferase assays are reported as in Fig. 1D.