ADRENOCORTICOTROPIN-RESISTANT MUTANTS OF THE Y1 ADRENAL-CELL LINE FAIL TO EXPRESS THE ADRENOCORTICOTROPIN RECEPTOR

This report examines the basis for adrenocorticotropin (ACTH) resistan ce in two mutant clones (Y6 and OS3) derived from the ACTH-responsive Y1 mouse adrenocortical tumor cell line. These two mutants were origin ally characterized by their failure to respond to ACTH with increased adenylyl cyclase activity and as a consequence were resistant to the s teroidogenic effects of the hormone. We now demonstrate that ACTH resi stance in the Y6 and OS3 mutants results from the failure to express t he gene encoding the ACTH receptor. Whereas parental Y1 cells express ACTH receptor transcripts at low,levels and are stimulated by ACTH or 8-bromo-cAMP to increase the accumulation of ACTH receptor transcripts approximately twofold, the Y6 and OS3 mutants do not express receptor transcripts either in the presence or absence of 8-bromo-cAMP. The ge ne encoding the ACTH receptor appears to be present in the Y6 and OS3 mutants, as determined by Southern blot hybridization analysis. Moreov er, in the Y6 mutant the ACTH receptor gene appears to be silenced by a modification that is reversed following the growth of the cells as t umors in mice. Clonal isolates of Y6 cells grown as tumors recover the ability to express ACTH receptor transcripts at low but detectable le vels and acquire the ability to respond to ACTH with increased adenyly l cyclase activity. Finally, Y6 and OS3 cells transformed with a gene encoding the mouse beta(2)-adrenergic receptor respond to the beta-adr energic agonist, isoproterenol, in a manner that is indistinguishable from the similarly transformed parent Y1 cell line. These latter resul ts demonstrate the functional integrity of the adenylyl cyclase system in the ACTH-resistant mutants and indicate that the failure to expres s ACTH receptor transcripts limits the responsiveness of these clones. (C) 1995 Wiley-Liss, Inc.