Mitochondrial metabolic reprogramming is important for many different processes like apoptosis and inflammation. NR4A1 is necessary for many different pro-apoptotic and inflammatory stimuli to induce mitochondrial metabolic reprogramming. However, despite the importance for small molecule ligands in the regulation of nuclear receptors, biological ligands regulating NR4A1 have not been identified. Here we report that glucose-1-phosphate (G1P), an intracellular metabolite from glycogen metabolism and glycolysis, is an NR4A1 ligand. In vitro, G1P specifically bound to NR4A1 ligand-binding domain (NR4A1LBD) and alters its oligomerization state. In cells, a synthetic cell-permeable G1P analog regulates transcriptional activity of NR4A1. This G1P analog suppressed mitochondrial respiration and induced apoptosis of H1299 lung adenocarcinoma cells in an NR4A1-dependent manner. This result suggests that G1P might be an endogenous ligand regulating NR4A1 and NR4A1-dependent mitochondrial metabolic reprogramming. It represents a rare case where a polar water-soluble intracellular metabolite modulates a nuclear receptor function.
Support or Funding Information
This work was supported by Johns Hopkins Provost’s Office Catalyst award, University of Kentucky start-up fund, and a National Institute of Health National Cancer Institute grant (R01CA168658) to Y.S.L. A.E.R. was supported in part by the University of Kentucky Undergraduate Summer Research Award. The University of Kentucky Flow Cytometry and Immune Monitoring Core Facility, the Oncogenomics Shared Resource Facility, the Bioinformatics Shared Resource Facility, and the Redox Metabolism Shared Resource Facility were supported by the Office of the Vice President for Research, the Markey Cancer Center, and a National Cancer Institute Center Core Support Grant (NIH P30 CA177558).
This work was supported by Johns Hopkins Provostamp;rsquo;s Office Catalyst award, University of Kentucky start-up fund, and a National Institute of Health National Cancer Institute grant (R01CA168658) to Y.S.L. A.E.R. was supported in part by the University of Kentucky Undergraduate Summer Research Award.amp;nbsp;The University of Kentucky Flow Cytometry and Immune Monitoring Core Facility, the Oncogenomics Shared Resource Facility, the Bioinformatics Shared Resource Facility, and the Redox Metabolism Shared Resource Facility wereamp;nbsp; supported by the Office of the Vice President for Research, the Markey Cancer Center, and a National Cancer Institute Center Core Support Grant (NIH P30 CA177558).amp;nbsp;
Fig. 1 | G1P binds to NR4A1LBD. a. ESI-MS spectra showing that an ion whose m/z was 283.0195 (+1 charged) was enriched with NR4A1LBD but not with NR4A3LBD. b. Trp fluorescence spectra of NR4A1LBD (0.5 mM) in the presence of varying amount of G1P. c. NR4A1LBD Trp fluorescence emission intensity at different concentrations of G1P and G6P. d. A simulation result showing the binding of G1P to the structure of NR4A1LBD. e. T608V and R576Q mutations suppresses the binding of G1P.; Fig. 2 | Cell-permeable G1P analog regulates NR4A1-dependent gene expression. a. G1CH2PPOM2 and G1CH2P, a G1P analog. b-c. Trp fluorescence emission spectra of NR4A1LBD showing the binding of G1CH2P to NR4A1LBD. d-e. Effect of G1CH2PPOM2 on the NR4A1-dependent reporter gene expression. f. RNA-Seq results showing genes differentially expressed by G1CH2PPOM2 in scrambled-KD and NR4A1-KD cells. g. Western blot result showing that G1CH2PPOM2 induced NR4A1-dependent expression of c-Fos.
