A region in the steroid binding domain determines formation of the non-DNA-binding, 9 S glucocorticoid receptor complex

• Published in: The Journal of biological chemistry 1988-01, Vol.263 (1), p.267-273
• Main Authors: Pratt, W B, Jolly, D J, Pratt, D V, Hollenberg, S M, Giguere, V, Cadepond, F M, Schweizer-Groyer, G, Catelli, M G, Evans, R M, Baulieu, E E
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Biological and medical sciences; Biotechnology; Cell Line; Cell receptors; Cell structures and functions; DNA - metabolism; Fundamental and applied biological sciences. Psychology; Genetic engineering; Genetic technics; glucocorticoid; Humans; Macromolecular Substances; Methods. Procedures. Technologies; Molecular and cellular biology; Molecular cloning; Molecular Sequence Data; Mutation; Plasmids; Receptors, Glucocorticoid - genetics; Receptors, Glucocorticoid - metabolism; Species Specificity; steroid; Transfection; Triamcinolone Acetonide - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)57388-4

This work was initiated to determine if a specific region of the glucocorticoid receptor determines the formation of the inactive (i.e. non-DNA-binding) 9 S form of the receptor recovered in cytosol preparations. It is known that the murine glucocorticoid receptor of the nti phenotype, which consists of only the carboxyl-terminal 40-kDa peptide containing the DNA-binding and steroid-binding domains separated by a short linker region, is recovered in hypotonic lysates as a 9 S heteromeric complex (Gehring, U., and Arndt, H. (1985) FEBS Lett. 179, 138-142). To further localize the domain required for formation of the 9 S complex, we have determined the sedimentation coefficients of receptors produced in COS-7 cells transfected with several mutants of the human glucocorticoid receptor gene. Deletion of the DNA-binding domain results in a 9 S complex that is somewhat less stable than the wild type receptor during sucrose gradient centrifugation. Deletion of the linker region yields a molybdate-stabilized 9 S complex, but deletion of the entire steroid-binding domain or internal deletion of the amino-terminal two-thirds of this domain yields receptors that are constitutive transcriptional activators and are present in cytosol only in the 4 S form. Taken together, these observations demonstrate that the steroid-binding domain contains the features required for formation of the 9 S heteromeric complex, and they are consistent with the proposal that the steroid-binding domain normally represses receptor function.