Fig. 5: Mitochondria DNA damage accumulation and DNA repair defects can be rescued by the correction of FUS mutations or targeted Lig1 expression. | Nature Communications

Fig. 5: Mitochondria DNA damage accumulation and DNA repair defects can be rescued by the correction of FUS mutations or targeted Lig1 expression.

From: FUS unveiled in mitochondrial DNA repair and targeted ligase-1 expression rescues repair-defects in FUS-linked motor neuron disease

Fig. 5

a IF of endogenous FUS localization in control and patient derived motor neurons along with isogenic mutation corrected cells, IF was performed by anti-FUS antibody. DAPI staining indicates nucleus. Scale bar = 10 µm, the experiment was performed in three independent repetitions. b IB of endogenous FUS, HSP60, Tom20, and PCNA in FUS WT, FUS P525L and isogenic FUS L525P motor neurons, the data are presented as mean ± s.e.m. from three individual experiments, with the individual data points shown. c Human DNA ligase 1 (Lig1) localizing in mitochondria (FLAG-MTS) was generated with n-terminal FLAG and mitochondrial localization signal (MTS) from Cytochrome c oxidase subunit 8 (COX8) gene, IB showing Lig1expression in nucleus and mitochondria in patient derived fibroblasts, the experiment was performed in three independent repetitions. d In vitro nick ligation activity assay performed using patient derived NPSC mitochondrial extracts with MTS-Lig1 expression, the data are presented as mean ± s.e.m. from three individual experiments, with the individual data points shown. e LA-PCR-based DNA damage repair kinetic analysis. Genome DNA extracted from control and patient derived NPSC with FUS WT, FUS P525L and FUS P525L MTS-lig1 at indicated time points after release from exposure to GO (100 ng/ml) for 1 h. Amplification products analyzed by agarose gel electrophoresis and pico green-based quantitation represented, the LA-PCR data are presented as mean ± s.e.m. from three individual experiments, with the individual data points shown. f microplate reader-based analysis of TMRM signal intensity, comparison of control, FUS P525L and FUS P525L cells expressing MTS-Lig1. The cells were treated with GO (100 ng/ml) for 1 h and released for 2 h post 1 h treatment. the data are presented as mean ± s.e.m. from four individual experiments, with the individual data points shown. All statistical analysis were performed using two-sided student-t test in graph pad prism software. Source data are provided as a Source Data file.

Back to article page