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ACTH-receptor deficient mutants of the Y1 mouse adrenocortical tumor cell line
B P Schimmer1, W K Kwan, J Tsao
1Banting and Best Department of Medical Research, University of Toronto, Ontario, Canada.
Abstract:
Two mutant clones (Y6 and OS3) derived from the ACTH-responsive Y1 mouse adrenocortical tumor cell line fail to respond to ACTH with increased adenylyl cyclase activity and, as a consequence, are resistant to the steroidogenic effects of the hormone. As determined from Northern blot and RNase protection assays, ACTH resistance in these mutants results from the failure to accumulate ACTH receptor transcripts. The ACTH receptor gene appears to be present in these mutants as determined by Southern blot hybridization analysis and can be activated following the growth of the mutant cells as tumors in mice, suggesting that the ACTH receptor gene is modified in a reversible manner. When mutant cells are transformed with a gene encoding the mouse beta 2-adrenergic receptor they respond to beta-adrenergic agonists with increased adenylyl cyclase activity in a manner that is indistinguishable from a similarly transformed parent Y1 cell line. These results suggest that the adenylyl cyclase system in the mutants is otherwise intact and that the failure to express ACTH receptor transcripts limits the responsiveness of these clones to the hormone.
Insights
Two mouse adrenocortical tumor cell clones resistant to ACTH lack ACTH receptor transcripts. This ACTH receptor gene defect is reversible, suggesting a modified gene expression underlies hormone resistance.
Area of Science:
- Endocrinology
- Molecular Biology
- Cell Biology
Background:
- Y1 mouse adrenocortical tumor cells are responsive to ACTH.
- Mutant clones Y6 and OS3 exhibit resistance to ACTH.
- This resistance is characterized by a lack of adenylyl cyclase activation and steroidogenesis.
Purpose of the Study:
- To investigate the molecular basis of ACTH resistance in Y1 mouse adrenocortical tumor cell mutants.
- To determine if the ACTH receptor gene is present and functional in these resistant clones.
Main Methods:
- Northern blot and RNase protection assays to analyze ACTH receptor transcript levels.
- Southern blot hybridization to assess the presence of the ACTH receptor gene.
- Cellular transformation with a beta 2-adrenergic receptor gene to assess adenylyl cyclase system integrity.
Main Results:
- Mutant clones Y6 and OS3 failed to accumulate ACTH receptor transcripts.
- The ACTH receptor gene was detected in mutants, suggesting a reversible modification.
- Transformed mutants responded normally to beta-adrenergic agonists, indicating an intact adenylyl cyclase system.
Conclusions:
- ACTH resistance in Y6 and OS3 clones is due to a failure in ACTH receptor transcript accumulation.
- The ACTH receptor gene modification is reversible.
- The adenylyl cyclase system remains functional, with transcript deficiency limiting hormone responsiveness.