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Differential Regulation of Glucocorticoid and Progesterone Receptor Subcellular Localization by Tetratricopeptide Repeat Domain Proteins

Banerjee, Ananya
http://rave.ohiolink.edu/etdc/view?acc_num=mco1177535048

Abstract Details

2007, Doctor of Philosophy in Medical Sciences (Ph.D.), University of Toledo, College of Graduate Studies.
Steroid-receptor associated immunophilins not only act in a coordinated manner to establish and maintain receptor conformation optimal for signaling but also play a critical role in determining receptor localization. To demonstrate TPR preference in steroid receptor localization, we chose a mouse fibroblast cell line, which expresses high levels of endogenous GR, and a Chinese hamster ovary, derived cell line, WCL2, that over-expresses mouse GR localized in the nucleus in the unliganded condition. Immunofluorescence and fractionation studies showed that although GR is predominantly nuclear in the WCL2 cells, it is not tightly bound to this compartment and is released into the cytosolic fraction upon cell lysis. Comparing the differences in the equilibrium levels of FKBP51 and FKBP52 between L929 and WCL2 not only revealed a very cell-type specific expression pattern but also hinted at the possibility of differential recruitment. Analysis of heterocomplex composition identified FKBP52 as the major TPR in WCL2 cells while FKBP51 was preferentially recruited into GR complexes from L929 cells. Having demonstrated the TPR preferences of GR in both cell types, we over-expressed FLAG-tagged FKBP51 and FKBP52 to determine if receptor subcellular localization could be altered by changing the equilibrium levels of these proteins. Receptor heterocomplexes in WCL2 showed a greater cytoplasmic distribution resulting from the recruitment of FLAG-tagged FKBP51, while distribution of GR in L929 cells remained unaffected by the distribution of flag-tagged FKBP52. To assess if the subcellular localization of other steroid receptors as well hinged on their preference towards specific TPR proteins, GFP fluorescence and co-immunoprecipitation assays were performed on receptorless COS-1 cells expressing either GFP-tagged wild or chimeric receptor constructs of GR and PR-B. Preference of GR towards FKBP51 remained unaltered while PR-B complexes mostly recruited FKBP52. Similar studies performed on chimeric receptor constructs identified a dual role of the ligand-binding domain (LBD), which is involved in controlling TPR specificity of receptor as well as subcellular localization. Finally, by selectively deleting specific TPR proteins, we show that loss of FKBP51 had a minor impact on localization of both receptors; loss of FKBP52 had a dramatic impact on PR localization while absence of PP5 did not alter localization of either receptor sub-type. Taken together these results show that steroid receptor preference for specific TPR controls overall subcellular localization although this can be overcome by equilibrium levels of cell-type specific expression.
Edwin Sanchez (Advisor)
123 p.
Biology, Molecular
Glucocorticoid; Progesterone; Dexamethasone; Receptor; Cyclophillin; Tetratricopeptide

Recommended Citations

Citations

  • Banerjee, A. (2007). Differential Regulation of Glucocorticoid and Progesterone Receptor Subcellular Localization by Tetratricopeptide Repeat Domain Proteins [Doctoral dissertation, University of Toledo]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=mco1177535048

    APA Style (7th edition)

  • Banerjee, Ananya. Differential Regulation of Glucocorticoid and Progesterone Receptor Subcellular Localization by Tetratricopeptide Repeat Domain Proteins. 2007. University of Toledo, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=mco1177535048.

    MLA Style (8th edition)

  • Banerjee, Ananya. "Differential Regulation of Glucocorticoid and Progesterone Receptor Subcellular Localization by Tetratricopeptide Repeat Domain Proteins." Doctoral dissertation, University of Toledo, 2007. http://rave.ohiolink.edu/etdc/view?acc_num=mco1177535048

    Chicago Manual of Style (17th edition)

mco1177535048
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© 2007, all rights reserved.
This open access ETD is published by University of Toledo Health Science Campus and OhioLINK.