Glucocorticoid receptor structure and function in an adrenocorticotropin-secreting small cell lung cancer

• Published in: Molecular endocrinology (Baltimore, Md.) Vol. 9; no. 9; pp. 1193 - 1201
• Main Authors: Gaitan, D, DeBold, C R, Turney, M K, Zhou, P, Orth, D N, Kovacs, W J
• Format: Journal Article
• Language: English
• Published: United States 01.09.1995
• Subjects: Adrenocorticotropic Hormone - secretion; Animals; Base Sequence; Blotting, Northern; Carcinoma, Small Cell - secretion; DNA, Complementary - chemistry; Humans; Lung Neoplasms - secretion; Mice; Molecular Sequence Data; Receptors, Glucocorticoid - chemistry; Receptors, Glucocorticoid - genetics; Receptors, Glucocorticoid - physiology; RNA, Messenger - analysis; Signal Transduction; Transfection; Tumor Cells, Cultured
• ISSN: 0888-8809; 1944-9917
• DOI: 10.1210/me.9.9.1193

ACTH secretion by tumors of nonpituitary origin is characteristically resistant to negative feedback regulation by glucocorticoids. One possible mechanism for the phenomenon could be a structural defect in the intracellular glucocorticoid receptor (GR). We studied the GR in DMS-79 cells derived from a human ACTH-secreting small cell lung cancer. Compared with control cells, DMS-79 cells were found to have greatly diminished GR ligand-binding activity and immunoreactive 94-kilodalton (kDa) GR content. Northern blot analysis revealed expression of GR transcripts that appeared to be slightly larger than those in control cells. A DMS-79 cell GR cDNA was cloned by reverse transcription/polymerase chain reaction amplification of mRNA using primers specific for full-length normal GR. The derived sequence of this full-length GR differed from the reported sequence by a single altered codon (G to A; Asn to Ser at codon 363) outside the steroid-binding domain. This N363S DMS-79 GR functioned normally to activate a target gene [mouse mammary tumor virus-chloramphenicol acetyl transferase (MMTV-CAT)] in transient transfection experiments in COS cells. Evidence for expression of a second type of GR mRNA was obtained by screening a DMS-79 cell cDNA library. This GR cDNA contained normal GR sequence up to nucleotide 2155, corresponding exactly to the end of exon 7 in the normal GR gene. The sequence appended to the GR sequences was not matched by any known sequence in DNA databases and included an in-frame termination codon after only 6 bases. The predicted truncated GR protein product (GR delta) has a mol wt of 73,740 and lacks most of the ligand-binding domain. Transient transfection of the GR delta form into COS cells did not reveal any dominant negative effect on the function of a cotransfected normal GR.