Quantitative proteomics reveals that dormancy-related proteins mediate the attenuation in mycobacterium strains

Although members of the <i>Mycobacterium tuberculosis</i> complex (MTBC) exhibit high similarity, they are characterized by differences with respect to virulence, immune response, and transmissibility. To understand the virulence of these bacteria and identify potential novel therapeutic targets, we systemically investigated the total cell protein contents of virulent H37Rv, attenuated H37Ra, and avirulent <i>M. bovis</i> BCG vaccine strains at the log and stationary phases, based on tandem mass tag (TMT) quantitative proteomics. Data analysis revealed that we obtained deep-coverage protein identification and high quantification. Although 272 genetic variations were reported in H37Ra and H37Rv, they showed very little expression difference in log and stationary phase. Quantitative comparison revealed H37Ra and H37Rv had significantly dysregulation in log phase (227) compared with stationary phase (61). While BCG and H37Rv, and BCG and H37Ra showed notable differences in stationary phase (1171 and 1124) with respect to log phase (381 and 414). In the log phase, similar patterns of protein abundance were observed between H37Ra and BCG, whereas a more similar expression pattern was observed between H37Rv and H37Ra in the stationary phase. Bioinformatic analysis revealed that the upregulated proteins detected for H37Rv and H37Ra in log phase were virulence-related factors. In both log and stationary phases, the dysregulated proteins detected for BCG, which have also been identified as <i>M. tuberculosis</i> response proteins under dormancy conditions. We accordingly describe the proteomic profiles of H37Rv, H37Ra, and BCG, which we believe will potentially provide a better understanding of H37Rv pathogenesis, H37Ra attenuation, and BCG immuno protection.

尽管结核分枝杆菌复合群（Mycobacterium tuberculosis complex, MTBC）各成员间同源性极高，但在毒力、免疫应答与传播能力方面存在显著差异。为解析此类细菌的毒力机制并筛选潜在新型治疗靶点，本研究基于串联质量标签（tandem mass tag, TMT）定量蛋白质组学技术，系统检测了强毒株H37Rv、减毒株H37Ra以及无毒株牛分枝杆菌（*M. bovis*）BCG疫苗株在对数生长期与稳定生长期的全细胞蛋白表达谱。数据分析结果显示，本研究实现了高覆盖度的蛋白质鉴定与高精度定量。尽管已有研究报道H37Ra与H37Rv之间存在272处遗传变异，但二者在对数生长期与稳定生长期的蛋白表达差异极小。定量对比分析显示，相较于稳定生长期（61个差异表达蛋白），H37Ra与H37Rv在对数生长期的差异表达蛋白达227个，差异更为显著。而BCG分别与H37Rv、H37Ra的差异表达蛋白数量在稳定生长期分别为1171、1124个，显著高于对数生长期的381、414个。在对数生长期，H37Ra与BCG的蛋白丰度模式较为相似；而在稳定生长期，H37Rv与H37Ra的表达模式则更为接近。生物信息学分析表明，H37Rv与H37Ra在对数生长期上调的蛋白均为毒力相关因子。本研究检测到的BCG差异表达蛋白，均被证实为结核分枝杆菌在休眠状态下的应答蛋白。本研究系统阐明了H37Rv、H37Ra与BCG的蛋白质组学特征，我们认为该结果将有助于更深入地理解H37Rv的致病机制、H37Ra的减毒机制以及BCG的免疫保护作用。