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The Co-chaperones FKBP51 and PP5 Control Nuclear Receptor Phosphorylation and Adipogenesis

Stechschulte, Lance A

Abstract Details

2013, Doctor of Philosophy (PhD), University of Toledo, College of Medicine.
FK506-binding protein 51 (FKBP51), a steroid receptor co-chaperone, is a known negative regulator of glucocorticoid receptor (GR) activity. In this work, we show that FKBP51 is also a co-chaperone to peroxisome proliferator-activated receptor-γ (PPARγ) and it is essential for PPARγ activity. We investigate the mechanism by which FKBP51 can regulate GR and PPARγ. In receptor-less COS-7 cells transfected with GR and PPARγ, over-expression of FKBP51 reduced GR activity at a minimal GRE-luciferase reporter, while increasing PPARγ activity at a minimal PPRE-reporter. Conversely, use of FKBP51-deficient mouse embryonic fibroblast cells (FKBP51-KO MEFs) showed elevated GR but reduced PPARγ activities at both reporter and endogenous genes, compared to wild-type (WT) MEFs. Because phosphorylation is known to exert a similar pattern of reciprocal modulation of GR and PPARγ, the contribution of FKBP51 to the phosphorylation states of each receptor was assessed. Knockdown of FKBP51 via shRNA in 3T3-L1 pre-adipocytes showed elevated phosphorylation of PPARγ at serine 112, a phospho-residue known to inhibit receptor activity. In FKBP51-KO MEF cells, elevated phosphorylation of GR at serines 220 and 234 was observed, phospho-residues known to promote receptor activity. Because FKBP51 lacks a phosphatase domain indirect regulation of phosphorylation was assessed. FKBP51 was recently discovered to chaperone the Akt-specific phosphatase PHLPP, Akt signaling was investigated in FKBP51-KO cells. Our data revealed elevated basal and insulin-induced Akt activation in the absence of FKBP51, as well as elevated activation of p38 mitogen-activated protein kinases (MAPK), a downstream target of Akt that directly phosphorylates both GR and PPARγ, causing activation and inhibition, respectively. Inhibition of p38 MAPK with PD169316 reversed the effects of FKBP51 deficiency on both GR and PPARγ. Loss of FKBP51 caused a redistribution of PPARγ from the cytoplasm to the nucleus, as previously shown for GR. A model is therefore proposed in which FKBP51 loss reciprocally regulates GR and PPARγ via two complementary mechanisms: activation of Akt-p38 MAPK mediated phosphorylation and redistribution of the receptors to the nucleus for direct targeting by p38 MAPK. In mouse 3T3-L1 preadipocyte cells, levels of steroid receptor co-chaperones known to interact with nuclear receptors, PP5 and FKBP51 were elevated in the differentiated state, while expression of FKBP52 and Cyp40 were decreased. Importantly, knockdown of FKBP51 via shRNA in 3T3-L1 pre-adipocytes showed resistance to differentiation with a reduction in lipid accumulation and adipose specific gene markers. FKBP51-KO cells were highly resistant to differentiation, showing almost no intracellular lipid accumulation and reduction in adipose specific gene markers. This phenotype was completely reversed in FKBP51-KO cells following either reintroduction of FKBP51 or treatment with PD169316, a selective p38 MAPK inhibitor. Protein Phosphatase 5 (PP5), a second steroid receptor co-chaperone, is a known negative regulator of GR activity and positive regulator of PPARγ activity. PP5 was previously shown to regulate GR and PPARγ activity by controlling nuclear receptor phosphorylation (Hinds, Stechschulte et al. 2011). In this work, we observed PP5 mutant animals had reduced body fat and an increase in lean mass as measured by NMR. They had fasting hypoglycaemia, improved glucose tolerance and insulin sensitivity compared to WT littermate controls. They also had higher energy expenditure compared to WT animals during indirect calorimetry measurement. Taken as a whole, these results suggest that FKBP51 and PP5 may play a key role in regulating different branches of metabolism by altering receptor functions. Therefore, designing drugs that target FKBP51 and PP5, rather than the receptors themselves, might provide an opportunity to control many features of metabolic syndrome, atherosclerosis, dyslipidemia, obesity, type 2 diabetes, and osteoporosis with little or no side effects.
Edwin Sanchez, Ph.D. (Committee Chair)
Sonia Najjar, Ph.D. (Committee Member)
Beata Lecka-Czernik, Ph.D. (Committee Member)
Manohar Ratnam, Ph.D. (Committee Member)
Yatrik Shah, Ph.D. (Committee Member)
147 p.

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Citations

  • Stechschulte, L. A. (2013). The Co-chaperones FKBP51 and PP5 Control Nuclear Receptor Phosphorylation and Adipogenesis [Doctoral dissertation, University of Toledo]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=mco1370871316

    APA Style (7th edition)

  • Stechschulte, Lance. The Co-chaperones FKBP51 and PP5 Control Nuclear Receptor Phosphorylation and Adipogenesis. 2013. University of Toledo, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=mco1370871316.

    MLA Style (8th edition)

  • Stechschulte, Lance. "The Co-chaperones FKBP51 and PP5 Control Nuclear Receptor Phosphorylation and Adipogenesis." Doctoral dissertation, University of Toledo, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=mco1370871316

    Chicago Manual of Style (17th edition)