Evaluation of receptor function in ACTH-responsive and ACTH-insensitive adrenal tumor cells

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Abstract
Two variant cell lines (Y6 and OS3), derived from the ACTH-sensitive mouse adrenocortical tumor clone Y1, are defective in the ACTH-sensitive adenylate cyclase system. The nature of the defects in Y6 and OS3 cells was characterized using ACTH1-10, ACTH4-10 and cholera toxin. In Y1 cells, ACTH1-39, ACTH1-10, and ACTH4-10 stimulated steroidogenesis to the same maximum level with Kd'' values of 5 .times. 10-11 M, 5 .times. 10-7 M and 10-4 M, respectively. ACTH1-10 (0.4 mM) and ACTH4-10 (3.2 mM) increased the accumulation of cyclic[c]AMP in Y1 cells 2-3-fold. Cholera toxin increased steroidogenesis and cAMP accumulation in Y1 cells with Kd'' values of 0.4 ng/ml and 9 ng/ml respectively. Y6 and OS3 cells responded to added cholera toxin with increased cAMP accumulation and increased steroidogenesis but did not respond to ACTH1-39, ACTH1-10 or ACTH4-10 at concentrations effective in Y1 cells. Y6 and OS3 cells may be defective in a process or component that links the principal binding regions of the ACTH receptor to the catalytic subunit of the adenylate cyclase system. The interactions of ACTH with the principal binding regions of the ACTH receptor were studied by analysis of binding of radioactive, iodinated ACTH1-24. ACTH binding showed low affinity, high capacity and no target-tissue specificity, and was considered not to be useful in evaluating the integrity of the ACTH receptor.