Abstract：
Mycobacterium tuberculosis (Mtb) successfully thrives in the host by adjusting its metabolism and manipulating the host environment. In this study, we investigated the role of Rv0547c, a protein that carries mitochondria-targeting sequence (MTS), in mycobacterial persistence. We show that Rv0547c is a functional oxidoreductase that targets host-cell mitochondria. Interestingly, the localization of Rv0547c to mitochondria was independent of the predicted MTS but depended on specific arginine residues at the N- and C-terminals. As compared to the mitochondria-localization defective mutant, Rv0547c-2SDM, wild-type Rv0547c increased mitochondrial membrane fluidity and spare respiratory capacity. To comprehend the possible reason, comparative lipidomics was performed that revealed a reduced variability of long-chain and very long-chain fatty acids as well as altered levels of phosphatidylcholine and phosphatidylinositol class of lipids upon expression of Rv0547c, explaining the increased membrane fluidity. Additionally, the over representation of propionate metabolism and β-oxidation intermediates in Rv0547c-targeted mitochondrial fractions indicated altered fatty acid metabolism, which corroborated with changes in oxygen consumption rate (OCR) upon etomoxir treatment in HEK293T cells transiently expressing Rv0547c, resulting in enhanced mitochondrial fatty acid oxidation capacity. Furthermore, Mycobacterium smegmatis over expressing Rv0547c showed increased persistence during infection of THP-1 macrophages, which correlated with its increased expression in Mtb during oxidative and nutrient starvation stresses. This study identified for the first time an Mtb protein that alters mitochondrial metabolism and aids in survival in host macrophages by altering fatty acid metabolism to its benefit and, at the same time increases mitochondrial spare respiratory capacity to mitigate infection stresses and maintain cell viability.
摘要：
结核分枝杆菌（Mtb）通过调节其代谢和操纵宿主环境而成功地在宿主中繁殖。在这项研究中,我们调查了Rv0547c的作用，一种携带线粒体靶向序列（MTS）的蛋白质，分枝杆菌的持久性。我们证明Rv0547c是一种靶向宿主细胞线粒体的功能性氧化还原酶。有趣的是,Rv0547c在线粒体上的定位与预测的MTS无关，但取决于N端和C端的特定精氨酸残基。与线粒体定位缺陷突变体相比，Rv0547c-2SDM,野生型Rv0547c增加线粒体膜流动性和备用呼吸能力。为了理解可能的原因，进行了比较脂质组学，揭示了长链和非常长链脂肪酸的变异性降低，以及在Rv0547c表达时脂质的磷脂酰胆碱和磷脂酰肌醇类别的水平改变，解释膜流动性增加。此外,Rv0547c靶向线粒体部分中丙酸代谢和β-氧化中间体的过度表达表明脂肪酸代谢改变，这证实了在瞬时表达Rv0547c的HEK293T细胞中，依托莫昔尔处理后耗氧率（OCR）的变化，导致线粒体脂肪酸氧化能力增强。此外，过表达Rv0547c的耻垢分枝杆菌在THP-1巨噬细胞感染期间显示出持久性增加，这与氧化和营养饥饿胁迫期间其在Mtb中的表达增加有关。这项研究首次确定了一种Mtb蛋白，它通过改变脂肪酸代谢来改变线粒体代谢并有助于宿主巨噬细胞的存活，同时增加线粒体备用呼吸能力，以减轻感染压力并维持细胞活力。
