Dysfunctional glucocorticoid receptor with a single point mutation ablates the CCAAT/enhancer binding protein‐dependent growth suppression response in a steroid‐resistant rat hepatoma cell variant

Abstract

We used glucocorticoid-resistant and -sensitive hepatoma cell variants to characterize the mechanism of hepatoma cell resistance to the growth inhibitory effects of glucocorticoids. BDS1 hepatoma cells express transcriptionally active glucocorticoid receptors and undergo a stringent G1 cell cycle arrest in response to glucocorticoids that is dependent on the induced expression of the CCAAT/enhancer binding protein alpha (C/EBPalpha) transcription factor. In contrast, EDR1 hepatoma cells, which express normal levels of glucocorticoid receptors, fail to growth arrest or express C/EBPalpha when treated with glucocorticoids. Ectopic expression of wild-type rat glucocorticoid receptors into EDR1 cells restored the growth suppression response, suggesting a defect in the EDR1 receptor. DNA sequence analysis revealed a single point mutation causing a cysteine-to-tyrosine substitution at amino acid position 457 (C457Y-GR) in the zinc finger region of the glucocorticoid receptor that mediates both receptor-DNA and receptor-protein interactions. Glucocorticoid activation of the alpha1-acid glycoprotein (AGP) promoter, a liver acute-phase response gene, requires receptor-DNA binding as well as an interaction with C/EBPalpha. In contrast to the wild-type glucocorticoid receptor, ectopic expression of C/EBPalpha in EDR1 cells, or coexpression of C/EBPalpha along with the C457Y-GR into receptor-deficient EDR3 cells was required to partially restore glucocorticoid responsiveness of the AGP promoter by the EDR1 glucocorticoid receptor. Constitutive expression of the wild-type glucocorticoid receptor, but not the C457Y-GR mutant, was sufficient to restore the glucocorticoid growth suppression response to receptor-deficient EDR3 cells. Thus, we have identified a glucocorticoid-resistant hepatoma cell variant with a single point mutation in the zinc finger region of the glucocorticoid receptor gene that ablates the glucocorticoid growth suppression response and attenuates transcriptional activation of the AGP promoter.

CCAAT enhancer binding protein β and hepatocyte nuclear factor 3β are necessary and sufficient to mediate dexamethasone-induced up-regulation of alpha2HS-glycoprotein/fetuin-A gene expression

“…Unlike several studies of genomic effects of dexamethasone (Cram et al 1998, Ramos et al 1999, Rüdiger et al 2002 we did not observe a contribution of C/EBP-in dexamethasone-mediated stimulation of the mouse Ahsg gene, because an antibody against C/EBP-was unable to alter transcription factor binding at the Ahsg SRU.…”

Krüppel-like Factors Regulate the Lama1 Gene Encoding the Laminin α1 Chain

“…Indeed, C/EBP␤ as well as C/EBP␦ expression and binding have been shown to be rapidly induced by GC and to be involved in the responsiveness to GC in the IEC-6 intestinal epithelial cell line (43). In addition, interactions between C/EBP proteins and glucocorticoid receptors are involved in the transcriptional effect induced by steroids (59). However, mutation or deletion of the C/EBP␤ site did not allow us to demonstrate such an interaction for the DX-induced transcriptional activation of Lama1.…”

Role of Glucocorticoid Receptor in Serosa-Involved Gastric Carcinoma After Gastrectomy

“…22 Thereafter, GR was found in leukemia, melanoma cells, and various types of cancer cells. [6][7][8][9][10][11][12][13]19 However, the clinical significance of GR in various malignancies differs. In human hepatocellular carcinoma, we found that the prognosis of GR-positive patients was poorer than that of GR-negative patients.…”

“…[6][7][8][9][10][11][12][13] Although the action of glucocorticoid on the cancer cells is through GR, its effects differ in different cancers. For example, in human adult acute lymphoblastic leukemia, the prognosis of the patients whose leukemic cellular GR level can be reduced by administration of glucocorticoid is better than those whose leukemic cellular GR level cannot be reduced by glucocorticoid.…”

“…[2][3][4][5] On the other hand, glucocorticoid has been claimed to enhance or limit the progression of various cancers. [6][7][8][9][10][11][12][13] The biologic effects have been reported to be through the glucocorticoid receptor (GR) in the cancer cell. [10][11][12][13] More than 15 years ago, GR was also found in the cytosol of gastric carcinoma.…”