Table_6_The TbD1 Locus Mediates a Hypoxia-Induced Copper Response in Mycobacterium bovis.XLSX

The Mycobacterium tuberculosis complex (MTBC) contains the causative agents of tuberculosis (TB) in mammals. The archetypal members of the MTBC, Mycobacterium tuberculosis and Mycobacterium bovis, cause human tuberculosis and bovine tuberculosis, respectively. Although M. tuberculosis and M. bovis share over 99.9% genome identity, they show distinct host adaptation for humans and animals; hence, while the molecular basis of host adaptation is encoded in their genomes, the mechanistic basis of host tropism is still unclear. Exploration of the in vitro phenotypic consequences of known genetic difference between M. bovis and M. tuberculosis offers one route to explore genotype–phenotype links that may play a role in host adaptation. The TbD1 (“Mycobacterium tuberculosis deletion 1 region”) locus encompasses the mmpS6 and mmpL6 genes. TbD1 is absent in M. tuberculosis “modern” lineages (Lineages 2, 3, and 4) but present in “ancestral” M. tuberculosis (Lineages 1 and 7), Mycobacterium africanum lineages (Lineages 5 and 6), newly identified M. tuberculosis lineages (Lineages 8 and 9), and animal adapted strains, such as M. bovis. The function of TbD1 has previously been investigated in M. tuberculosis, where conflicting data has emerged on the role of TbD1 in sensitivity to oxidative stress, while the underlying mechanistic basis of such a phenotype is unclear. In this study, we aimed to shed further light on the role of the TbD1 locus by exploring its function in M. bovis. Toward this, we constructed an M. bovis TbD1 knockout (ΔTbD1) strain and conducted comparative transcriptomics to define global gene expression profiles of M. bovis wild-type (WT) and the ΔTbD1 strains under in vitro culture conditions (rolling and standing cultures). This analysis revealed differential induction of a hypoxia-driven copper response in WT and ΔTbD1 strains. In vitro phenotypic assays demonstrated that the deletion of TbD1 sensitized M. bovis to H2O2 and hypoxia-specific copper toxicity. Our study provides new information on the function of the TbD1 locus in M. bovis and its role in stress responses in the MTBC.

结核分枝杆菌复合群（Mycobacterium tuberculosis complex，简称MTBC）包含哺乳动物结核病的致病菌。MTBC的典型成员，即结核分枝杆菌（Mycobacterium tuberculosis）和牛分枝杆菌（Mycobacterium bovis），分别引起人类和牛的结核病。尽管M. tuberculosis和M. bovis的基因组同源性超过99.9%，但它们在人类和动物宿主适应性方面表现出显著的差异；因此，尽管宿主适应性的分子基础编码在其基因组中，但宿主嗜性的机制基础尚不明确。探究已知遗传差异在牛分枝杆菌（M. bovis）和结核分枝杆菌（M. tuberculosis）之间的体外表型后果，为探索可能在宿主适应性中发挥作用的基因型-表型联系提供了一条途径。TbD1（“结核分枝杆菌删除1区域”）基因座包含mmpS6和mmpL6基因。TbD1在M. tuberculosis的“现代”谱系（谱系2、3和4）中不存在，但在“祖先”M. tuberculosis（谱系1和7）、非洲分枝杆菌（Mycobacterium africanum）谱系（谱系5和6）、新发现的M. tuberculosis谱系（谱系8和9）以及动物适应性菌株，如M. bovis中存在。TbD1的功能先前已在M. tuberculosis中进行了研究，关于TbD1在氧化应激敏感性方面的作用出现了矛盾的数据，而这种现象的潜在机制基础尚不明确。在本研究中，我们旨在通过探索TbD1基因座在M. bovis中的功能，进一步阐明其在MTBC应激反应中的作用。为此，我们构建了M. bovis TbD1敲除（ΔTbD1）菌株，并进行了比较转录组学分析，以定义M. bovis野生型（WT）和ΔTbD1菌株在体外培养条件（滚动和静置培养）下的全局基因表达谱。该分析揭示了WT和ΔTbD1菌株中缺氧驱动的铜反应的差异诱导。体外表型分析表明，TbD1的缺失使M. bovis对H2O2和缺氧特异性铜毒性敏感。我们的研究为TbD1基因座在M. bovis中的功能及其在MTBC应激反应中的作用提供了新的见解。