Department of Microbiology, College of Life Sciences, Nanjing Agricultural University, Nanjing, 210095, Jiangsu, People's Republic of China.
Appl Microbiol Biotechnol. 2023 Feb;107(4):1361-1371. doi: 10.1007/s00253-022-12357-4. Epub 2023 Jan 13.
Mitochondrial pyruvate carriers (MPCs), located in the inner membrane of mitochondria, are essential carriers for pyruvate to enter mitochondria. MPCs regulate a wide range of intracellular metabolic processes, such as glycolysis, the tricarboxylic acid cycle (TCA cycle), fatty acid metabolism, and amino acid metabolism. However, the metabolic regulation of MPCs in macrofungi is poorly studied. We studied the role of MPCs in Ganoderma lucidum (GlMPC) on ganoderic acid (GA) biosynthesis regulation in G. lucidum. In this study, we found that the mitochondrial/cytoplasmic ratio of pyruvate was downregulated about 75% in GlMPC1- and GlMPC2-silenced transformants compared with wild type (WT). In addition, the GA content was 17.72 mg/g and increased by approximately 50% in GlMPC1- and GlMPC2-silenced transformants compared with WT. By assaying the expression levels of three key enzymes and the enzyme activities of isocitrate dehydrogenase (IDH) and α-ketoglutarate dehydrogenase (α-KGDH) of the TCA cycle in GlMPC1- and GlMPC2-silenced transformants, it was found that the decrease in GlMPCs activity did not significantly downregulate the TCA cycle rate, and the enzyme activity of IDH increased by 44% compared with WT. We then verified that fatty acid β-oxidation (FAO) supplements the TCA cycle by detecting the expression levels of key enzymes involved in FAO. The results showed that compared with WT, the GA content was 1.14 mg/g and reduced by approximately 40% in co-silenced transformants. KEY POINTS: • GlMPCs affects the distribution of pyruvate between mitochondria and the cytoplasm. • Acetyl-CoA produced by FAO maintains the TCA cycle. • Acetyl-CoA produced by FAO promotes the accumulation of GA.
线粒体丙酮酸载体（MPC）位于线粒体的内膜上，是丙酮酸进入线粒体的必需载体。MPC 调节广泛的细胞内代谢过程，如糖酵解、三羧酸循环（TCA 循环）、脂肪酸代谢和氨基酸代谢。然而，宏观真菌中 MPC 的代谢调节研究甚少。我们研究了 Ganoderma lucidum（GlMPC）中 MPC 对灵芝酸（GA）生物合成调节的作用。在这项研究中，我们发现与野生型（WT）相比，GlMPC1 和 GlMPC2 沉默转化体的线粒体/细胞质丙酮酸比约下调 75%。此外，GlMPC1 和 GlMPC2 沉默转化体中的 GA 含量为 17.72mg/g，比 WT 增加约 50%。通过测定 GlMPC1 和 GlMPC2 沉默转化体中三个关键酶的表达水平和三羧酸循环中的异柠檬酸脱氢酶（IDH）和α-酮戊二酸脱氢酶（α-KGDH）的酶活性，发现 GlMPCs 活性的降低并没有显著下调 TCA 循环速率，并且 IDH 的酶活性比 WT 增加了 44%。然后，我们通过检测参与 FAO 的关键酶的表达水平来验证 FAO 是否补充 TCA 循环。结果表明，与 WT 相比，共沉默转化体中的 GA 含量为 1.14mg/g，减少了约 40%。关键点：• GlMPCs 影响丙酮酸在线粒体和细胞质之间的分布。• FAO 产生的乙酰 CoA 维持 TCA 循环。• FAO 产生的乙酰 CoA 促进 GA 的积累。
