Isolation of glucocorticoid-unresponsive rat hepatoma cells by fluorescence-activated cell sorting.
We have used a mouse mammary tumor virus (MMTV)-infected rat hepatoma cell line as a model system for studying glucocorticoid action. These cells induce tyrosine aminotransferase and MMTV in response to the synthetic glucocorticoid, dexamethasone. The major viral antigen, a glycoprotein of 52,000 daltons (gp52), appears on the surface of infected cells in amounts which reflect the cytoplasmic content of viral RNA. Using an anti-gp52 antiserum and a fluorescence-activated cell sorter (FACS), we have selected variants which display low levels of pg52 in the presence of the hormone. Multiple cycles of enrichment for cells that fluoresce weakly in the presence of hormone have generated a population which fails to produce a detectable increase in cell surface gp52 in response to dexamethasone. This population of nonresponders and a number of independent clones derived from this population were analyzed for their ability to induce gp52 and TAT and for these presence of glucocorticoid receptors. All nonresponder clones exhibited little or no induction of either glucocorticoid-inducible marker. Two of the clones contained reduced levels of glucocrticoid receptor, while the remainder of the clones showed no detectable specific hormone binding. These results provide genetic evidence that a single class of glucocorticoid receptors is involved in the induction of both MMTV and TAT in HTC cells.
D000818:Animals; D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D002469:Cell Separation / Q000379:methods; D003907:Dexamethasone / Q000494:pharmacology*; D004790:Enzyme Induction; D008113:Liver Neoplasms; D008324:Mammary Tumor Virus, Mouse; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D011987:Receptors, Steroid / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D014444:Tyrosine Transaminase / Q000096:biosynthesis*; D014764:Viral Proteins / Q000096:biosynthesis*
D011965:Receptors, Glucocorticoid; D011987:Receptors, Steroid; D014764:Viral Proteins; D003907:Dexamethasone; D014444:Tyrosine Transaminase