Linking epigenetic dysregulation, mitochondrial impairment, and metabolic dysfunction in SBMA motor neurons
Naemeh Pourshafie, … , Christopher Grunseich, Kenneth H. Fischbeck
Naemeh Pourshafie, … , Christopher Grunseich, Kenneth H. Fischbeck
Published July 9, 2020
Citation Information: JCI Insight. 2020;5(13):e136539. https://doi.org/10.1172/jci.insight.136539.
View: Text | PDF
Research Article Genetics Neuroscience

Linking epigenetic dysregulation, mitochondrial impairment, and metabolic dysfunction in SBMA motor neurons

Abstract

Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular disorder caused by a polyglutamine expansion in the androgen receptor (AR). Using gene expression analysis and ChIP sequencing, we mapped transcriptional changes in genetically engineered patient stem cell–derived motor neurons. We found that transcriptional dysregulation in SBMA can occur through AR-mediated histone modification. We detected reduced histone acetylation, along with decreased expression of genes encoding compensatory metabolic proteins and reduced substrate availability for mitochondrial function. Furthermore, we found that pyruvate supplementation corrected this deficiency and improved mitochondrial function and SBMA motor neuron viability. We propose that epigenetic dysregulation of metabolic genes contributes to reduced mitochondrial ATP production. Our results show a molecular link between altered epigenetic regulation and mitochondrial metabolism that contributes to neurodegeneration.

Authors

Naemeh Pourshafie, Ester Masati, Eric Bunker, Alec R. Nickolls, Parisorn Thepmankorn, Kory Johnson, Xia Feng, Tyler Ekins, Christopher Grunseich, Kenneth H. Fischbeck

×

Figure 1

iMNs differentiated from SBMA iPSCs show increased cellular stress and cell death.

Options: View larger image (or click on image) Download as PowerPoint
iMNs differentiated from SBMA iPSCs show increased cellular stress and c...
(A) Representative images of iMNs (6 dpi) expressing HB9, ISL1, and TUJ1. Scale bars: 75 μm. (B and C) Percentage of HB9+/TUJ1+ (B) and ISL1+/TUJ1+ (C), assessed by immunostaining. n = 4–5 per cell line. (D–F) Bioenergetic extracellular flux analysis (Seahorse assay) on iMNs normalized to total cellular protein concentration. (D) Rate of ATP production during oxidative phosphorylation (MitoATP production rate). (E) Rate of ATP production in the glycolytic pathway (glycoATP production rate). (F) The sum of the glycolytic and mitochondrial ATP production rates (total ATP production rate). n = 16–38 wells/group. (G) Ratiometric pseudocolor images of GoATEAM expressed in iMNs. ATP sensors were introduced into iMNs using lentivirus 4 days before taking the first image. (H) Comparison of orange/green fluorescence emission ratio of GoATEAM at different time points. The ratio was calculated from fluorescence images. Plates were seeded at the same density, and live images were taken from the same plate over time. On average, 30 images per cell line were used for calculating the ratio at each time point. (I) Representative images of dying cells with less plasma membrane integrity were detected with a florescent stain in real time. NucGreen dead 488 (green), iMNs expressing the hNIL-mCherry plasmid (red), and DAPI (blue). (J) Comparison of GFP/DAPI emission ratio of NucGreen dead at different time points. The ratio was calculated from fluorescence images. n = 4–6 per group. All experiments were performed on N = 3 SBMA, N = 3 control, and N = 3 AR-KO. Error bars show mean ± SE; *P < 0.05, ***P < 0.001, ****P < 0.0001. One-way ANOVA followed by Bonferroni’s multiple comparisons test. iMNs were treated with 10 nM DHT. Scale bars: 25 μm (G) and 40 μm (I).