Defects in CYB5A and CYB5B impact sterol-C4 oxidation in cholesterol biosynthesis and demonstrate regulatory roles of dimethyl sterols

• Published in: Cell reports (Cambridge) 2024-11, Vol.43 (11), p.114912, Article 114912
• Main Authors: Ma, Mei-Yan, Deng, Gang, Zhu, Wen-Zhuo, Sun, Ming, Jiang, Lu-Yi, Li, Wei-Hui, Liu, Yuan-Bin, Guo, Lin, Song, Bao-Liang, Zhao, Xiaolu
• Format: Article
• Language: English
• Subjects: Animals; Cholesterol - biosynthesis; Cholesterol - metabolism; CP: Metabolism; CYB5A/CYB5B; Cytochromes b5 - genetics; Cytochromes b5 - metabolism; HeLa Cells; Humans; Liver - metabolism; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Oxidation-Reduction; SREBP and PPARγ pathway; sterol C4-demethylation; Sterols - metabolism; T-MAS/dihydro-T-MAS
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2024.114912

Cytochrome b5 (CYB5) is a hemoprotein crucial for electron transfer to oxygenases. Although microsomal CYB5A is required for sterol C4-demethylation in vitro, cholesterol biosynthesis remains intact in Cyb5a knockout mice. Here, we show that knockout of mitochondrial CYB5B, rather than CYB5A, blocks cholesterol biosynthesis at the sterol-C4 oxidation step in HeLa cells, causing an accumulation of testis meiosis-activating sterol (T-MAS) and dihydro-T-MAS. Surprisingly, liver-specific Cyb5b knockout (L-Cyb5b−/−) mice exhibit normal cholesterol metabolism. Further knockdown of Cyb5a in L-Cyb5b−/− (L-Cyb5b−/−/short hairpin [sh]Cyb5a) mice leads to a marked accumulation of T-MAS and dihydro-T-MAS, indicating that either CYB5A or CYB5B is required for sterol C4-demethylation. The L-Cyb5b−/−/shCyb5a mice are largely normal, with lower sterol regulatory element-binding protein (SREBP)-target gene expression during refeeding and higher liver triglyceride levels while fasting, as T-MAS and dihydro-T-MAS inhibit the SREBP pathway and activate the PPARγ pathway. In summary, CYB5A and CYB5B compensate for sterol C4-demethylation, and T-MAS and dihydro-T-MAS can modulate the SREBP and PPARγ pathways. [Display omitted] •CYB5B, not CYB5A, is required for sterol C4-demethylation in HeLa cells•L-Cyb5b−/− mice exhibit normal cholesterol metabolism•CYB5A and CYB5B functionally compensate for sterol C4-demethylation•T-MAS and dihydro-T-MAS inhibit the SREBP pathway while activating PPARγ Cytochrome b5 (CYB5) is a hemoprotein crucial for electron transfer to oxygenases. Ma et al. show that CYB5A and CYB5B functionally compensate for sterol C4-demethylation in cholesterol biosynthesis. Depletion of CYB5A and CYB5B causes the accumulation of SC4MOL substrates T-MAS and dihydro-T-MAS, which inhibit the SREBP pathway and activate PPARγ.