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Mitochondrial FAD shortage in SLC25A32 deficiency affects folate-mediated one-carbon metabolism

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Abstract

The SLC25A32 dysfunction is associated with neural tube defects (NTDs) and exercise intolerance, but very little is known about disease-specific mechanisms due to a paucity of animal models. Here, we generated homozygous (Slc25a32Y174C/Y174C and Slc25a32K235R/K235R) and compound heterozygous (Slc25a32Y174C/K235R) knock-in mice by mimicking the missense mutations identified from our patient. A homozygous knock-out (Slc25a32−/−) mouse was also generated. The Slc25a32K235R/K235R and Slc25a32Y174C/K235R mice presented with mild motor impairment and recapitulated the biochemical disturbances of the patient. While Slc25a32−/− mice die in utero with NTDs. None of the Slc25a32 mutations hindered the mitochondrial uptake of folate. Instead, the mitochondrial uptake of flavin adenine dinucleotide (FAD) was specifically blocked by Slc25a32Y174C/K235R, Slc25a32K235R/K235R, and Slc25a32−/− mutations. A positive correlation between SLC25A32 dysfunction and flavoenzyme deficiency was observed. Besides the flavoenzymes involved in fatty acid β-oxidation and amino acid metabolism being impaired, Slc25a32−/− embryos also had a subunit of glycine cleavage system—dihydrolipoamide dehydrogenase damaged, resulting in glycine accumulation and glycine derived-formate reduction, which further disturbed folate-mediated one-carbon metabolism, leading to 5-methyltetrahydrofolate shortage and other folate intermediates accumulation. Maternal formate supplementation increased the 5-methyltetrahydrofolate levels and ameliorated the NTDs in Slc25a32−/− embryos. The Slc25a32K235R/K235R and Slc25a32Y174C/K235R mice had no glycine accumulation, but had another formate donor—dimethylglycine accumulated and formate deficiency. Meanwhile, they suffered from the absence of all folate intermediates in mitochondria. Formate supplementation increased the folate amounts, but this effect was not restricted to the Slc25a32 mutant mice only. In summary, we established novel animal models, which enabled us to understand the function of SLC25A32 better and to elucidate the role of SLC25A32 dysfunction in human disease development and progression.

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The datasets generated during or analyzed during the current study are all available in the manuscript.

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Acknowledgements

We warmly thank the patient and her parents for participating in this study. We also thank Zhu-Yuan Xu (Account Manager, ProteinSimple) and Zu-Sen Weng (FAS, ProteinSimple) for their help in the Western blot assays, Zhe Li (Ph.D, School of Pharmaceutical Sciences, Sun Yat-sen University) for his help in the homology modeling analysis, and Prof. Ya-Ping Tang (Guangzhou Women and Children’s Medical Center) and Prof. Guo-Jun Shi (The third affiliated hospital, Sun-Yat Sen University) for their valuable suggestions in preparing the manuscript.

Funding

This study was funded by the National Natural Science Foundation of China (81802125 to M-Z P., 81700755 to Y-X S., and 81873661 to L L.), and Guangzhou Women and Children’s Medical Center/Guangzhou Institute of Pediatrics (IP-2018-024 to M-Z P.). The funding sources are not involved with the preparation of the article.

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  1. Min-Zhi Peng, Yong-Xian Shao and Xiu-Zhen Li: Co-first authors.

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  1. Department of Genetics and Endocrinology, Guangzhou Women and Children’s Medical Center, the Affiliated Hospital of Guangzhou Medical University, 9 Jinsui Road, Guangzhou, China

    Min-Zhi Peng, Yong-Xian Shao, Xiu-Zhen Li, Kang-Di Zhang, Yan-Na Cai, Yun-Ting Lin, Min-Yan Jiang, Zong-Cai Liu, Xue-Ying Su, Wen Zhang, Xiao-Ling Jiang & Li Liu

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Contributions

Study concept and design were undertaken by M-Z P, Y-X S, and L L. Data acquisition and analysis were undertaken by M-Z P, Y-X S, X-Z L, K-D Z, Y-N C, Y-T L, M-Y J, Z-C L, X-Y S, W Z, and X-L J. The manuscript was written by M-Z P. and revised by L L, Y-X S, and X-L J.

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Correspondence to Wen Zhang, Xiao-Ling Jiang or Li Liu.

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This study was performed in line with the principles of the Declaration of Helsinki. The ethics committee of Guangzhou Women and Children’s Medical Center authorized this study, and the Institutional Animal Care and Use Committee of Guangzhou Medical University approved the animal studies. Informed consent has been obtained from the patient and her guardian.

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Peng, MZ., Shao, YX., Li, XZ. et al. Mitochondrial FAD shortage in SLC25A32 deficiency affects folate-mediated one-carbon metabolism. Cell. Mol. Life Sci. 79, 375 (2022). https://doi.org/10.1007/s00018-022-04404-0

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