The biochemical basis of mitochondrial dysfunction in Zellweger Spectrum Disorder (2021)
Figure 6

Figure 6. Overexpression of ATAD1 rescues mitochondrial respiration and metabolism.

(A) Bioenergetic profile of human fibroblast cell lines. OCR (pmol/min/norm.unit) for cWT, cWT+ATAD1, PEX3-/-, PEX3-/-+ATAD1, cWT-ATAD1-/-, and PEX3-/- atad1-/- cells plotted against time (XF96e Extracellular Flux Analyzer, Mito-Stress-Test). 1 μM Oligomycin A, 0.15 μM FCCP, and 1 μM rotenone + 1 μM antimycin A (final concentrations) were sequentially delivered to the XF96e assay medium through injection ports in the sensor cartridge. The graph is the compilation of two independent assays, error bars show mean +/- s.d. (n=4, biological replicates).

(B) Bar graph representation of basal respiration measured in (B) for cWT, cWT+ATAD1 , PEX3-/-, PEX3-/-+ATAD1, cWT-ATAD1 , and PEX3-/--atad1 cells. Error bars show mean +/- s.d. (n=4, biological replicates). Statistical significance was calculated using the Welch's test, p-values are indicated as: ns (not significant), ** p ≤ 0.01, *** p ≤ 0.001.

(C) Bar graph representation of maximal respiration measured in (B) for cWT, cWT+ATAD1 , PEX3-/-, PEX3-/-+ATAD1, cWT-ATAD1 and PEX3-/- -atad1 cells. Error bars show mean +/- s.d. (n=4, biological replicates). Statistical significance was calculated using the Welch's test, p-values are indicated as: ns (not significant), * p ≤ 0.05, ** p ≤ 0.01, **** p ≤ 0.0001.

(D) Bar graph representation of calculated respiratory spare capacity (uncoupled respiration minus basal respiration) measured in (B) for cWT, cWT+ATAD1 , PEX3-/-, PEX3-/-+ATAD1, cWT-ATAD1, and PEX3-/--atad1 cells. Error bars show mean +/- s.d. (n=4, biological replicates). Statistical significance was calculated using the Welch's test, p-values are indicated as: ns (not significant), * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001.

(E) Scatter plot representation of cardiolipin lipidomics. The average (n = 4, biological replicates) normalized peak intensity of each of the 11 detected cardiolipin species, log10 pareto-scaled is visualized. Statistical significance was calculated using the Welch's test, p-values are indicated as: ns (not significant), * p ≤ 0.05, ** p ≤ 0.01. The horizontal lines represent the mean peak intesity of all 11 caridolipin species in each cell line.

­­(F) Scatter plot representation of a subpopulation of the phosphoethanolamine (PE) lipidomics. The average (n = 4, biological replicates) normalized peak intensity of each of the 20 phosphoethanolamine (PE) species (decreased in PEX3-/-), log10 pareto-scaled, is visualized. Statistical significance was calculated using the Welch's test, p-values are indicated as: ns (not significant), * p ≤ 0.05, *** p ≤ 0.001, **** p ≤ 0.0001. The horizontal lines represent the mean peak intesity of all 20 PE species in each cell line.

(G) Scatter plot representation of ether-phospholipid lipidomics. The average (n = 4, biological replicates) normalized peak intensity of each of the 21 detected ether-phospholipid (plasmalogen) species, log10 pareto-scaled, is visualized. Statistical significance was calculated using the Welch's test, p-values are indicated as: ns (not significant), *** p ≤ 0.001, **** p ≤ 0.0001. The horizontal lines represent the mean peak intesity of all 21 plasmalogen species in each cell line.

Expanded View Figure Legends