Figure 1From: Translation of MMTV Gag requires nuclear events involving splicing motifs in addition to the viral Rem protein and RmREGag constructs produce protein in vitro but not in mammalian cells. A) Schematic representation of the three gag expression constructs: Mtv-1 gag, HBRV gag and SM gag. CMV = human cytomegalovirus immediate early promoter; ORF = open reading frame; BGH polyA = bovine growth hormone polyadenylation site. B) HEK 293T cells and C) MEF cells were transfected with the indicated gag plasmids. Gag protein production was assayed 24 hr post-transfection by metabolic labeling with35S methionine (1 hr pulse) followed by immunoprecipitation with rabbit polyclonal anti-MMTV CA sera and an antibody to β-catenin to control for cellular equivalents. Pr77gag, Gag precursor (77 KDa) and β-catenin (98 KDa) are indicated. D) The three gag constructs were transcribed and translated in vitro in a rabbit reticulocyte lysate. A T7 promoter-driven β-galactosidase gene that produces a 110 KDa product was included as a system control (+C). E) Western immunoblot of in vitro translated Gag with MMTV anti-CA sera. In this case, the β-galactosidase reaction serves as a negative control for the antibody (-C).Back to article page