PGE1 and PGA1 bind to Nurr1 and activate its transcriptional function

Abstract

The orphan nuclear receptor Nurr1 is critical for the development, maintenance and protection of midbrain dopaminergic (mDA) neurons. Here we show that prostaglandin E1 (PGE1) and its dehydrated metabolite, PGA1, directly interact with the ligand-binding domain (LBD) of Nurr1 and stimulate its transcriptional function. We also report the crystallographic structure of Nurr1-LBD bound to PGA1 at 2.05 Å resolution. PGA1 couples covalently to Nurr1-LBD by forming a Michael adduct with Cys566, and induces notable conformational changes, including a 21° shift of the activation function-2 helix (H12) away from the protein core. Furthermore, PGE1/PGA1 exhibit neuroprotective effects in a Nurr1-dependent manner, prominently enhance expression of Nurr1 target genes in mDA neurons and improve motor deficits in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned mouse models of Parkinson’s disease. Based on these results, we propose that PGE1/PGA1 represent native ligands of Nurr1 and can exert neuroprotective effects on mDA neurons, via activation of Nurr1’s transcriptional function.

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Fig. 1: Identification of PGE1 and 8-iso-PGE1 as Nurr1 activators from brain tissue extracts.
Fig. 2: Crystal structure of PGA1-bound Nurr1-LBD and its molecular and functional analyses.
Fig. 3: Structural comparison of apo and PGA1-bound Nurr1-LBD.
Fig. 4: Direct binding of PGE1/PGA1 to Nurr1-LBD or EP2.
Fig. 5: Neuroprotective effect of PGE1/PGA1 is dependent on Nurr1.
Fig. 6: Neuroprotective effects of PGE1 and PGA1 in the MPTP-induced PD mouse model.

Data availability

The PDB accession code for the coordinates and structure factors of Nurr1-LBD in complex with PGA1 is PDB 5Y41. Source data for Figs. 13 and Extended Data Figs. 19 are presented with the paper.

References

  1. 1.

    Mangelsdorf, D. J. et al. The nuclear receptor superfamily: the second decade. Cell 83, 835–839 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. 2.

    Evans, R. M. & Mangelsdorf, D. J. Nuclear receptors, RXR, and the big bang. Cell 157, 255–266 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Kliewer, S. A., Lehmann, J. M. & Willson, T. M. Orphan nuclear receptors: shifting endocrinology into reverse. Science 284, 757–760 (1999).

    CAS  PubMed  Google Scholar 

  4. 4.

    Kurakula, K., Koenis, D. S., van Tiel, C. M. & de Vries, C. J. NR4A nuclear receptors are orphans but not lonesome. Biochim Biophys. Acta 1843, 2543–2555 (2014).

    CAS  PubMed  Google Scholar 

  5. 5.

    Pearen, M. A. & Muscat, G. E. Minireview: nuclear hormone receptor 4A signaling: implications for metabolic disease. Mol. Endocrinol. 24, 1891–1903 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Zetterstrom, R. H. et al. Dopamine neuron agenesis in Nurr1-deficient mice. Science 276, 248–250 (1997).

    CAS  PubMed  Google Scholar 

  7. 7.

    Kadkhodaei, B. et al. Nurr1 is required for maintenance of maturing and adult midbrain dopamine neurons. J. Neurosci. 29, 15923–15932 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  8. 8.

    Saijo, K. et al. A Nurr1/CoREST pathway in microglia and astrocytes protects dopaminergic neurons from inflammation-induced death. Cell 137, 47–59 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Chu, Y. et al. Nurr1 in Parkinson’s disease and related disorders. J. Comp. Neurol. 494, 495–514 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. 10.

    Moran, L. B. et al. Analysis of alpha-synuclein, dopamine and parkin pathways in neuropathologically confirmed Parkinsonian nigra. Acta Neuropathol. 113, 253–263 (2007).

    CAS  PubMed  Google Scholar 

  11. 11.

    Decressac, M., Volakakis, N., Bjorklund, A. & Perlmann, T. NURR1 in Parkinson disease–from pathogenesis to therapeutic potential. Nat. Rev. Neurol. 9, 629–636 (2013).

    CAS  PubMed  Google Scholar 

  12. 12.

    Kim, C. H., Leblanc, P. & Kim, K. S. 4-amino-7-chloroquinoline derivatives for treating Parkinson’s disease: implications for drug discovery. Expert Opin. Drug Discov. 11, 337–341 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Kim, K. S. Toward neuroprotective treatments of Parkinson’s disease. Proc. Natl Acad. Sci. USA 114, 3795–3797 (2017).

    CAS  PubMed  Google Scholar 

  14. 14.

    Wang, Z. et al. Structure and function of Nurr1 identifies a class of ligand-independent nuclear receptors. Nature 423, 555–560 (2003).

    CAS  PubMed  Google Scholar 

  15. 15.

    Kagaya, S. et al. Prostaglandin A2 acts as a transactivator for NOR1 (NR4A3) within the nuclear receptor superfamily. Biol. Pharm. Bull. 28, 1603–1607 (2005).

    CAS  PubMed  Google Scholar 

  16. 16.

    Lakshmi, S. P., Reddy, A. T., Banno, A. & Reddy, R. C. Molecular, chemical, and structural characterization of prostaglandin A2 as a novel agonist for Nur77. Biochem. J. 476, 2757–2767 (2019).

    CAS  PubMed  Google Scholar 

  17. 17.

    Vinayavekhin, N. & Saghatelian, A. Discovery of a protein-metabolite interaction between unsaturated fatty acids and the nuclear receptor Nur77 using a metabolomics approach. J. Am. Chem. Soc. 133, 17168–17171 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. 18.

    de Vera, I. M. et al. Identification of a binding site for unsaturated fatty acids in the orphan nuclear receptor Nurr1. ACS Chem. Biol. 11, 1795–1799 (2016).

    PubMed  PubMed Central  Google Scholar 

  19. 19.

    de Vera, I. M. S. et al. Defining a canonical ligand-binding pocket in the orphan nuclear receptor Nurr1. Structure 27, 66–77 e65 (2019).

    PubMed  Google Scholar 

  20. 20.

    Chintharlapalli, S. et al. Activation of Nur77 by selected 1,1-bis(3′-indolyl)-1-(p-substituted phenyl)methanes induces apoptosis through nuclear pathways. J. Biol. Chem. 280, 24903–24914 (2005).

    CAS  PubMed  Google Scholar 

  21. 21.

    Zhan, Y. et al. Cytosporone B is an agonist for nuclear orphan receptor Nur77. Nat. Chem. Biol. 4, 548–556 (2008).

    CAS  PubMed  Google Scholar 

  22. 22.

    Kim, C. H. et al. Nuclear receptor Nurr1 agonists enhance its dual functions and improve behavioral deficits in an animal model of Parkinson’s disease. Proc. Natl Acad. Sci. USA 112, 8756–8761 (2015).

    CAS  PubMed  Google Scholar 

  23. 23.

    Andersen, N. H. Dehydration of prostaglandins: study by spectroscopic method. J. Lipid Res. 10, 320–325 (1969).

    CAS  PubMed  Google Scholar 

  24. 24.

    Itoh, T. et al. Structural basis for the activation of PPARgamma by oxidized fatty acids. Nat Struct. Mol. Biol. 15, 924–931 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Codina, A. et al. Identification of a novel co-regulator interaction surface on the ligand binding domain of Nurr1 using NMR footprinting. J. Biol. Chem. 279, 53338–53345 (2004).

    CAS  PubMed  Google Scholar 

  26. 26.

    Aarnisalo, P., Kim, C. H., Lee, J. W. & Perlmann, T. Defining requirements for heterodimerization between the retinoid X receptor and the orphan nuclear receptor Nurr1. J. Biol. Chem. 277, 35118–35123 (2002).

    CAS  PubMed  Google Scholar 

  27. 27.

    Zhan, Y. Y. et al. The orphan nuclear receptor Nur77 regulates LKB1 localization and activates AMPK. Nat. Chem. Biol. 8, 897–904 (2012).

    CAS  PubMed  Google Scholar 

  28. 28.

    Li, L. et al. Impeding the interaction between Nur77 and p38 reduces LPS-induced inflammation. Nat. Chem. Biol. 11, 339–346 (2015).

    CAS  PubMed  Google Scholar 

  29. 29.

    Wang, W. J. et al. Orphan nuclear receptor TR3 acts in autophagic cell death via mitochondrial signaling pathway. Nat. Chem. Biol. 10, 133–140 (2014).

    CAS  PubMed  Google Scholar 

  30. 30.

    Wang, W. J. et al. Induction of autophagic death in cancer cells by agonizing TR3 and attenuating Akt2 activity. Chem. Biol. 22, 1040–1051 (2015).

    CAS  PubMed  Google Scholar 

  31. 31.

    Furuyashiki, T. & Narumiya, S. Stress responses: the contribution of prostaglandin E(2) and its receptors. Nat. Rev. Endocrinol. 7, 163–175 (2011).

    CAS  PubMed  Google Scholar 

  32. 32.

    Carrasco, E., Casper, D. & Werner, P. PGE(2) receptor EP1 renders dopaminergic neurons selectively vulnerable to low-level oxidative stress and direct PGE(2) neurotoxicity. J. Neurosci. Res. 85, 3109–3117 (2007).

    CAS  PubMed  Google Scholar 

  33. 33.

    Parga, J. A. et al. Prostaglandin EP2 receptors mediate mesenchymal stromal cell-neuroprotective effects on dopaminergic neurons. Mol. Neurobiol. 55, 4763–4776 (2018).

    CAS  PubMed  Google Scholar 

  34. 34.

    Xu, H. et al. The MDM2-binding region in the transactivation domain of p53 also acts as a Bcl-X(L)-binding motif. Biochemistry 48, 12159–12168 (2009).

    CAS  PubMed  Google Scholar 

  35. 35.

    Becker, W., Bhattiprolu, K. C., Gubensak, N. & Zangger, K. Investigating protein-ligand interactions by solution nuclear magnetic resonance spectroscopy. Chem. Phys. Chem. 19, 895–906 (2018).

    CAS  PubMed  Google Scholar 

  36. 36.

    Choi, H. K., Won, L., Roback, J. D., Wainer, B. H. & Heller, A. Specific modulation of dopamine expression in neuronal hybrid cells by primary cells from different brain regions. Proc. Natl Acad. Sci. USA 89, 8943–8947 (1992).

    CAS  PubMed  Google Scholar 

  37. 37.

    Gao, L., Zhou, W., Symmes, B. & Freed, C. R. Re-cloning the N27 dopamine cell line to improve a cell culture model of Parkinson’s disease. PLoS ONE 11, e0160847 (2016).

    PubMed  PubMed Central  Google Scholar 

  38. 38.

    af Forselles, K. J. et al. In vitro and in vivo characterization of PF-04418948, a novel, potent and selective prostaglandin EP(2) receptor antagonist. Br. J. Pharmacol. 164, 1847–1856 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Bergstroem, S. & Samuelsson, B. Prostaglandins. Annu. Rev. Biochem. 34, 101–108 (1965).

    CAS  PubMed  Google Scholar 

  40. 40.

    Higdon, A., Diers, A. R., Oh, J. Y., Landar, A. & Darley-Usmar, V. M. Cell signalling by reactive lipid species: new concepts and molecular mechanisms. Biochem. J. 442, 453–464 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Qin, Z. H. et al. Prostaglandin A(1) protects striatal neurons against excitotoxic injury in rat striatum. J. Pharmacol. Exp. Ther. 297, 78–87 (2001).

    CAS  PubMed  Google Scholar 

  42. 42.

    Wang, X. et al. Prostaglandin A1 inhibits rotenone-induced apoptosis in SH-SY5Y cells. J. Neurochem. 83, 1094–1102 (2002).

    CAS  PubMed  Google Scholar 

  43. 43.

    Zhang, H. L. et al. Neuroprotective effects of prostaglandin A1 in animal models of focal ischemia. Brain Res. 1039, 203–206 (2005).

    CAS  PubMed  Google Scholar 

  44. 44.

    Bruning, J. M. et al. Covalent modification and regulation of the nuclear receptor nurr1 by a dopamine metabolite. Cell Chem. Biol. 26, 674–685 e676 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  45. 45.

    de Urquiza, A. M. et al. Docosahexaenoic acid, a ligand for the retinoid X receptor in mouse brain. Science 290, 2140–2144 (2000).

    PubMed  Google Scholar 

  46. 46.

    Janowski, B. A., Willy, P. J., Devi, T. R., Falck, J. R. & Mangelsdorf, D. J. An oxysterol signalling pathway mediated by the nuclear receptor LXR alpha. Nature 383, 728–731 (1996).

    CAS  PubMed  Google Scholar 

  47. 47.

    Powell, W. S. 15-Deoxy-delta12,14-PGJ2: endogenous PPARgamma ligand or minor eicosanoid degradation product? J. Clin. Invest. 112, 828–830 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Battye, T. G., Kontogiannis, L., Johnson, O., Powell, H. R. & Leslie, A. G. iMOSFLM: a new graphical interface for diffraction-image processing with MOSFLM. Acta Crystallogr. D. 67, 271–281 (2011).

    CAS  PubMed  Google Scholar 

  49. 49.

    Evans, P. Scaling and assessment of data quality. Acta Crystallogr. D. 62, 72–82 (2006).

    PubMed  Google Scholar 

  50. 50.

    Winn, M. D. et al. Overview of the CCP4 suite and current developments. Acta Crystallogr. D. 67, 235–242 (2011).

    CAS  PubMed  Google Scholar 

  51. 51.

    McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  52. 52.

    Murshudov, G. N. et al. REFMAC5 for the refinement of macromolecular crystal structures. Acta Crystallogr. D. 67, 355–367 (2011).

    CAS  PubMed  Google Scholar 

  53. 53.

    Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D. 60, 2126–2132 (2004).

    PubMed  Google Scholar 

  54. 54.

    Pettersen, E. F. et al. UCSF Chimera–a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004).

    CAS  PubMed  PubMed Central  Google Scholar 

  55. 55.

    Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D. 66, 213–221 (2010).

    CAS  PubMed  Google Scholar 

  56. 56.

    Waudby, C. A., Ramos, A., Cabrita, L. D. & Christodoulou, J. Two-dimensional NMR lineshape analysis. Sci. Rep. 6, 24826 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  57. 57.

    Leblanc, P. et al. Production of Nurr-1 specific polyclonal antibodies free of cross-reactivity against its close homologs, Nor1 and Nur77. J. Vis. Exp. 102, e52963 (2015).

    Google Scholar 

  58. 58.

    Kim, W. et al. miR-126 contributes to Parkinson’s disease by dysregulating the insulin-like growth factor/phosphoinositide 3-kinase signaling. Neurobiol. Aging 35, 1712–1721 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  59. 59.

    Chen, S. H., Oyarzabal, E. A. & Hong, J. S. Preparation of rodent primary cultures for neuron-glia, mixed glia, enriched microglia, and reconstituted cultures with microglia. Methods Mol. Biol. 1041, 231–240 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We thank various members of the molecular neurobiology laboratory past and present who participated in the project. In particular, we thank B.-S. Han, H.-Y. Jung, J. Lee and J. Sung Koh for technical assistance. We also acknowledge the contribution of the scientists and staff on the PXII and PXIII (Paul Scherrer Institute, Switzerland) beamlines for their expert assistance during crystal data collection. This work was supported by NIH grant nos. NS070577 and NS084869 (to K.-S.K.), NRF-2018M3A9B5023055 grant (to C.-H.K.), Ministry of Education Singapore AcRF Tier 2 Grant (no. ARC55/16) and Tang Tieng See Advancement Fund (to H.S.Y.), and National Medical Research Council, Singapore (grant no. TCR/013-NNI/2014; to K.L.L. and H.S.Y.).

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Authors

Contributions

K.-S.K. and H.S.Y. initiated and supervised the project. S.R., Y.J., C.H.K. and W.K. were responsible for the overall design and performance of experiments. S.R., H.T.T., H.Y., C.K., A.S., J.L., J.Y.Y., S.B., H.Y., C.K., X.L., G.G. and K.L.L. performed and analyzed structural studies. Y.J., C.H.K., W.K, J.J., B.S., M.F., Y.K., D.H., H.M.P., S.F.O. and C.H.L. performed and analyzed functional and biological studies. K.-S.K., H.S.Y., S.R., Y.J., C.H.K., W.K. and G.A.P. analyzed the data and wrote the paper. All authors contributed to the discussion and final approval of the paper.

Corresponding authors

Correspondence to Ho Sup Yoon or Kwang-Soo Kim.

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H.S.Y. is a nonexecutive director of Lifex Biolabs. The remaining authors have no competing interests to disclose.

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Supplementary information

Extended Data Fig. 1 Identification of PGE1 from brain tissue extract.

(a) Isolation of endogenous ligand candidates using a combination of boiling, acetone precipitation, and ultrafiltration (3,000 molecular weight cut-off). Fractions were monitored for their Nurr1-enhancing activity using a cell-based luciferase assay system. Nurr1-enhancing activity was unaffected by boiling and acetone precipitation. n = 3 independent experiments, Data are presented as mean ± s.d. (b) Following ultrafiltration, filtrates were fractionated by HPLC column C-18 and each fraction was assayed for Nurr1-activating activities. Fraction 5 contained the most activity and thus was used for the mass spectrometry (MS) analysis. n = 3 independent experiments, Data are presented as mean ± s.d. (c) Candidate compounds that are tested after identification by an ultra-performance liquid chromatography quadrupole time of flight mass spectrometry (UPLC–qTOF-MS/MS).

Extended Data Fig. 2 Direct binding of PGE1 and PGA1 to Nurr1-LBD.

(a–f) Molecular interaction of Nurr1-LBD with PGE1 (a and b) and PGA1 (c and d) studied using 2D HSQC NMR titration experiments with uniformly 15N-labeled Nurr1-LBD. (a) Close-up view of a section of the overlay of free Nurr1-LBD (red) and Nurr1-LBD with PGE1 (1:4, green; 1:10, blue), with residues showing chemical shift perturbations (with arrows) or intensity changes (boxed in red) labelled. (b) Selected residues were mapped on the crystal structure of Nurr1-LBD (PDB: 1OVL) in surface representation, with a close-up section (as inset) showing affected helices H4, H11 and H12, with amino acid residues indicated. (c) Close-up view of a section of the overlay of free Nurr1-LBD (red) and Nurr1-LBD with PGA1 (1:5, green; 1:10, blue), with residues showing chemical shift perturbations (with arrows) or intensity changes (boxed in red) labelled. Residues (Leu559, Gln571 and Thr595) showing additional peaks upon PGA1 incubation are marked with asterisks (*). This indicates that the PGE1 (a) and PGA1 (c) interaction with Nurr1-LBD matches the typical two-state binding model (P + L PL) and an induced-fit binding model (P + L PLopen → PLclosed), respectively. (d) Mapping of Nurr1-LBD residues perturbed in the presence of PGA1 reveals that both PGE1 (a and b) and PGA1 (c and d) recognize the same binding region on Nurr1-LBD, with maximum perturbation observed in helices H11 and H12. Residues showing chemical shifts and line broadening are coloured in purple while L410 is coloured in red (b and d), as its peak disappeared upon PGA1 binding. (e, f) PGA1 increases the transcriptional activity of Nurr1-based reporter constructs: Nurr1-LBD-dependent (e) and full-length Nurr1-dependent (f) transcriptional activities in SK-N-BE(2)C cells. n = 3 independent experiments, Data are presented as mean ± s.e.m.

Extended Data Fig. 3 Chemical shift perturbation plot of Nurr1-LBD upon PGE1 and PGA1 binding.

Chemical shift perturbation plot of Nurr1-LBD upon PGE1 (a) / PGA1 (b) binding (1:10 ratio) and their corresponding peak intensity plots (PGE1 (c) / PGA1 (d)) revealing residues with perturbed resonances and/or line broadening upon ligand binding. (*) denotes the peak belonging to L410 which disappeared upon PGA1 binding.

Extended Data Fig. 4 PGE1 conversion to PGA1 under crystal condition.

(a) The overlaid 2Fo-Fc (blue) and composite omit (pink) electron density maps contoured at 1σ cut-off confirming the conversion of PGE1 to PGA1, evident from the covalent bonding density with Cys566. (b) Mass spectrometry data of PGE1 incubated with Nurr1-LBD under crystallization buffer condition (100 mM MES, pH 5.5 and 200 mM MgCl2) confirming the conversion of PGE1 to PGA1, as revealed by the covalent complex molecular mass of 30,862 Da (Nurr1-LBD328–598 is 30,525 Da and PGA1 is 336.5 Da).

Extended Data Fig. 5 Crystal structure of PGA1-bound Nurr1-LBD and its molecular and functional analyses.

(a) Cartoon representation of Nurr1-LBD (blue) with PGA1 shown in sphere mode. (b) Interactions between PGA1 and Nurr1 residues (labelled) through hydrophobic contacts (grey broken lines) and hydrogen bonds (blue broken lines). Only chain B in the asymmetric unit are shown here, as the electron density for the PGA1 attached to this chain was complete. (c) PGJ2 and 15d-PGJ2 show no effect on the transcriptional activity of Nurr1-LBD. n = 3 independent experiments, Data are presented as mean ± s.e.m. (d) 15d-PGJ2 (3 μM), but not PGE1 (1 μM) or PGA1 (10 μM), induces the transcriptional activity of PPARγ-LBD.

Extended Data Fig. 6 Effects of mutations at Nurr1 residues interacting with the chain B (Arg515, His516, Arg563, Thr567).

(a), with the chain A (Phe443, Leu570, Ile573, Leu591) (b), and effects of mutations at the residue Cys566 (c) on PGA1 (10 μM)-induced transcriptional activation of Nurr1-LBD in SK-N-BE(2)C cells. n = 3 independent experiments, Data are presented as mean ± s.e.m.

Extended Data Fig. 7 Effects of EP2 agonists and antagonists on the transcriptional activity of Nurr1-LBD.

(a) The EP2 agonist, AH13205 activates Nurr1’s transcriptional activity, whereas EP3/EP4 agonists (Sulprostone and CAY10598) do not. (b) EP2 antagonist, PF-04418948 suppresses PGE1-induced transcriptional activation of Nurr1, whereas EP1/EP3/EP4 antagonists (SC-19220, L-798106, and L-161982) do not. (c) The synthetic PGE1 analogue misoprostol, activates Nurr1’s transcriptional activity in SK-N-BE(2)C cells. n = 3 independent experiments, Data are presented as mean ± s.e.m.

Extended Data Fig. 8 Protective effects of PGE1 and PGA1 against MPP+ in MN9D cells.

(a, b) Determination of protective effects of PGE1 and PGA1 in MN9D cells under MPP+-induced oxidative stress measured by MTT reduction. (a) Cells were treated with MPP+ (0–1000 µM) for 24 hrs. Cell viabilities assessed by MTT reduction assay show that treatment with 500 µM of MPP+ significantly induces 50% of cell death. (b) Pre-treatments with PGE1/PGA1 (24 hrs prior to MPP+ treatment) increase cell viability against the MPP+ induced oxidative stress in MN9D cells. *P < 0.05, **P < 0.01 compared to 0 µM; ###P < 0.001 compared to the absence of MPP+, unpaired two-tailed t-test; n = 3 independent samples per group. Data are mean ± s.e.m. (c, d) Protective effects of PGE1 and PGA1 measured by LDH release. (c) Cytotoxicity determined by LDH release assay also reveals that treatment with 500 µM of MPP+ significantly induces 50% of cell death in MN9D cells. (d) Similar to MTT reduction assay, pre-treatments with PGE1/PGA1 reduce cytotoxicity under the MPP+-induced oxidative stress. **P < 0.01, ***P < 0.001 compared to 0 µM; ###P < 0.001 compared to the absence of MPP+, unpaired two-tailed t-test; n = 3 independent samples per group. Data are mean ± s.e.m.

Extended Data Fig. 9 Effects of PGE1/PGA1 in the MPTP-induced reduction of DA levels.

The administration of PGE1/PGA1 significantly restores the MPTP-induced reduction of DA levels in the SN (a) and in the striatum (b). One-way ANOVA, Tukey’s post-hoc test; n = 5 per group. Data are mean ± s.e.m.

Extended Data Fig. 10 Mass spectrometry data between PGA1 and Nurr1-LBD under NMR condition.

Mass spectrometry data confirming the formation of the covalent bond between PGA1 (red line) with Nurr1-LBD356–598 (28.035 kDa), while PGE1 (blue dotted line) does not form such a covalent attachment under the NMR buffer conditions (20 mM sodium phosphate (pH 7.5) buffer containing 50 mM NaCl, 0.01% NaN3 in 90% H2O/10% D2O). The apo Nurr1-LBD356–598 (black line) (27.698 kDa) is shown for reference. The molecular weight of PGA1 is 336.5 Da. This also corroborates with the two-state binding and induced-fit model observed from NMR data (Extended Data Fig. 2a, c).

Supplementary information

Supplementary Information

Supplementary Figs. 1–18 and Tables 1–4.

Reporting Summary

Supplementary Video 1

PGA1 binding on Nurr1-LBD

Supplementary Video 2

Superposition of unbound over PGA1 bound Nurr1

Supplementary Video 3

Conformational changes induced by PGA1 binding

Source data

Source Data Fig. 1

Uncut Gel for Fig. 1g

Source Data Fig. 2

Uncut Gel for Fig. 2d

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Rajan, S., Jang, Y., Kim, C. et al. PGE1 and PGA1 bind to Nurr1 and activate its transcriptional function. Nat Chem Biol 16, 876–886 (2020). https://doi.org/10.1038/s41589-020-0553-6

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