Abstract
Analysis of the expression and transport of mouse mammary tumor virus (MMTV) glycoproteins in the viral infected rat hepatoma (HTC) cell line M1.54 has revealed a novel glucocorticoid-regulated trafficking pathway that controls the sorting and processing of cell surface MMTV glycoproteins. Complement cytolysis was used to recover M1.54-derived variants that fail to express a threshold level of cell surface MMTV glycoproteins. One complement selected variant, CR4, is selectively defective in the glucocorticoid-regulated sorting pathway while the expression of MMTV RNA is fully glucocorticoid responsive. Indirect immunofluorescence of intracellular and cell surface MMTV glycoproteins revealed that hormone induced CR4 displays an intracellular perinuclear staining of viral glycoproteins indicative of a block in the Golgi. 50 kb genomic fragments were prepared from DNA isolated from uninfected rat HTC hepatoma cells, ligated to the neomycin resistance gene and transfected into variant CR4 cells. DNA mediated complementation of CR4 was observed in 0.1% of neomycin resistant colonies screened for cell surface MMTV glycoproteins by indirect immunofluorescence. This phenotypic reversion was dependent upon glucocorticoid treatment. In a parallel approach, a truncated MMTV glycoprotein which lacks its transmembrane domain and cytoplasmic tail fails to be secreted and remains in an intracellular vesicle compartment in the presence of glucocorticoids. We conclude that rat hepatoma cells encode a glucocorticoid regulated trafficking (GRT) gene whose protein product acts in trans on specific substrates to mediate the sorting of cell surface glycoproteins.
Published Version
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