Fig. 7

Functional characterization of human OGDHL variants in zebrafish model. A Experimental approach for characterizing functionality of human OGDHL variants. Human OGDHL variant mRNAs were mixed with synthetic zebrafish-specific ogdhl single-guide RNAs (sgRNAs) and Cas9 protein and microinjected into one-cell stage embryos, followed by phenotypic evaluation at 3 dpf. Blue line indicates head size measurement approach. Red line indicates eye size measurement. Green arrowhead indicates heart edema. B, C Head and eye size measurements were calculated as a percentage difference compared to the mean value of the uninjected embryos. Subsequently, the calculated measurements of uninjected, Cas9-injected, and ogdhl F₀ embryos were compared. Additionally, the ogdhl F₀ co-injected with either WT or mutated OGDHL mRNA were compared to the ogdhl F₀ knockout embryos. Each dot represents one animal and the mean value of each group is indicated at the bottom of the respective bar in the figure. n = the number of embryos. Error bars = mean ± SD. Statistical significance was calculated by Brown–Forsthye and Welch’s ANOVA with Dunnett’s T3 multiple comparisons test: not significant (ns) p ≥ 0.05, *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. Significances for all rescue experiments are in comparison to ogdhl F₀. D The presence of the heart edema phenotype was calculated as a percentage of total embryos of each group. E The schematic of OGDHL truncating variants and the corresponding rescue experiment results