Glucocorticoids autoregulate their own glucocorticoid receptor (GR) mRNA, protein, and hormone-binding activity levels. In the mouse AtT-20 pituitary tumor cell line, the hormone coordinately regulates both GR and c-jun mRNA levels. Preliminary results indicate the involvement of primarily transcriptional components in the autoregulation of GR and c-jun mRNA. The first specific aim of this proposal is to determine the mechanism by which the GR and c-jun mRNA levels are coordinately regulated by glucocorticoid treatment. Preliminary studies in the mouse AtT-20 cell line have demonstrated an oscillatory regulation of both of these mRNAs, while studies on F9 teratocarcinoma and CEM-C7 leukemia cells have shown that both of these mRNAs are coordinately up-regulated when the cells are treated with retinoic acid and glucocorticoids, respectively. These observations will be extended in the nontumorigenic mouse L929 fibroblasts and in the human T-lymphoid cell line, CEM-C7. Confirmation of coordinate GR and c-jun gene expression will lead to experiments to determine (by nuclear run-on assays) if the transcription rates of both genes show corresponding coordinate regulation. Using an immunological analysis, the second specific aim will determine if the GR and Jun proteins form a heteromeric complex that results in transcriptional interference of AP-1 (Fos/Jun) activity. These experiments would be among the first to demonstrate in vivo cross-talk between these two intracellular signaling pathways at physiological levels of GR and Jun. The third specific aim will focus on the promoter elements in the c-jun and GR genes that are involved in the coordinate down-regulation of GR and c-jun mRNA and protein levels. The glucocorticoid response element (GRE) in the c-jun promoter will be mutated, as will the AP-1 sites in the c-jun and GR promoters. Cotransfection/ reporter gene expression experiments will then be used to determine if cross-talk, and hence coordinate regulation, has been abolished by these mutations. Intracellular Jun levels will be lowered using antisense approaches to directly determine if Jun is involved in GR gene regulation. In addition to focusing on the AP-1 sites in the promoters of these two genes, additional promoter analyses will be performed to insure that other potentially important cis-acting sequences are identified. Particular attention will be paid to the GR promoter, as no definitive functional studies have yet been performed to characterize this promoter. Important promoter elements involved in both the basal and hormone-regulated expression of the GR gene will thus be identified. These studies will reveal the molecular mechanism of basal and steroid- mediated alterations in GR and c-jun mRNA gene expression and the cross- talk that occurs between these two important intracellular signaling pathways. This could lead to novel therapeutic approaches for the treatment of steroid-hormone regulated cancers by modulating the functional glucocorticoid receptor and Jun status in the cell.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK047211-02
Application #
2146625
Study Section
Biochemical Endocrinology Study Section (BCE)
Project Start
1994-08-01
Project End
1998-07-31
Budget Start
1995-09-01
Budget End
1996-07-31
Support Year
2
Fiscal Year
1995
Total Cost
Indirect Cost
Name
Louisiana State University Hsc New Orleans
Department
Biochemistry
Type
Schools of Dentistry
DUNS #
782627814
City
New Orleans
State
LA
Country
United States
Zip Code
70112
Nunez, B Scott; Geng, Chuan-Dong; Pedersen, Kim Brint et al. (2005) Interaction between the interferon signaling pathway and the human glucocorticoid receptor gene 1A promoter. Endocrinology 146:1449-57
Geng, Chuan-dong; Pedersen, Kim Brint; Nunez, B Scott et al. (2005) Human glucocorticoid receptor alpha transcript splice variants with exon 2 deletions: evidence for tissue- and cell type-specific functions. Biochemistry 44:7395-405
Pedersen, Kim Brint; Geng, Chuan-dong; Vedeckis, Wayne V (2004) Three mechanisms are involved in glucocorticoid receptor autoregulation in a human T-lymphoblast cell line. Biochemistry 43:10851-8
Geng, Chuan-Dong; Vedeckis, Wayne V (2004) Steroid-responsive sequences in the human glucocorticoid receptor gene 1A promoter. Mol Endocrinol 18:912-24
Pedersen, Kim Brint; Vedeckis, Wayne V (2003) Quantification and glucocorticoid regulation of glucocorticoid receptor transcripts in two human leukemic cell lines. Biochemistry 42:10978-90
Nunez, B Scott; Vedeckis, Wayne V (2002) Characterization of promoter 1B in the human glucocorticoid receptor gene. Mol Cell Endocrinol 189:191-9
Breslin, M B; Geng, C D; Vedeckis, W V (2001) Multiple promoters exist in the human GR gene, one of which is activated by glucocorticoids. Mol Endocrinol 15:1381-95
Wagner, B L; Pollio, G; Giangrande, P et al. (1999) The novel progesterone receptor antagonists RTI 3021-012 and RTI 3021-022 exhibit complex glucocorticoid receptor antagonist activities: implications for the development of dissociated antiprogestins. Endocrinology 140:1449-58
Breslin, M B; Vedeckis, W V (1998) The human glucocorticoid receptor promoter upstream sequences contain binding sites for the ubiquitous transcription factor, Yin Yang 1. J Steroid Biochem Mol Biol 67:369-81
Wei, P; Inamdar, N; Vedeckis, W V (1998) Transrepression of c-jun gene expression by the glucocorticoid receptor requires both AP-1 sites in the c-jun promoter. Mol Endocrinol 12:1322-33

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