Table3_Host factor RBMX2 promotes epithelial cell apoptosis by downregulating APAF-1’s Retention Intron after Mycobacterium bovis infection.xls

The Mycobacterium tuberculosis variant bovis (M. bovis) is a highly pathogenic environmental microorganism that causes bovine tuberculosis (bTB), a significant zoonotic disease. Currently, “test and culling” is the primary measure for controlling bTB, but it has been proven to be inadequate in animals due to their high susceptibility to the pathogen. Selective breeding for increased host resistance to bTB to reduce its prevalence is feasible. In this study, we found a vital host-dependent factor, RBMX2, that can potentially promote M. bovis infection. By knocking RBMX2 out, we investigated its function during M. bovis infection. Through transcriptome sequencing and alternative splicing transcriptome sequencing, we concluded that after M. bovis infection, embryo bovine lung (EBL) cells were significantly enriched in RNA splicing associated with apoptosis compared with wild-type EBL cells. Through protein/molecular docking, molecular dynamics simulations, and real-time quantitative PCR, we demonstrated that RBMX2 promotes the apoptosis of epithelial cells by upregulating and binding to apoptotic peptidase activating factor 1 (APAF-1), resulting in the alternative splicing of APAF-1 as a retention intron. To our knowledge, this is the first report of M. bovis affecting host epithelial cell apoptosis by hijacking RBMX2 to promote the intron splicing of downstream APAF-1. These findings may represent a significant contribution to the development of novel TB prevention and control strategies.

牛分枝杆菌（Mycobacterium tuberculosis variant bovis, M. bovis）是一种高致病性环境病原微生物，可引发牛结核病（bovine tuberculosis, bTB）——一种危害严重的人畜共患病。当前，“检测与扑杀”是防控bTB的主要手段，但由于动物对该病原菌易感性极强，该措施已被证实效果有限。通过选择性育种提升宿主对bTB的抗性以降低其流行率，具备可行性。本研究中，我们发现了一种关键的宿主依赖因子RBMX2，其可在一定程度上促进M. bovis的感染过程。我们通过敲除RBMX2基因，探究了其在M. bovis感染过程中的功能。通过转录组测序与可变剪接转录组测序分析，我们发现：相较于野生型牛胚肺（embryo bovine lung, EBL）细胞，感染M. bovis后的EBL细胞在与细胞凋亡相关的RNA剪接通路中显著富集。通过蛋白质/分子对接、分子动力学模拟以及实时定量聚合酶链反应（real-time quantitative PCR）实验，我们证实：RBMX2可通过上调凋亡蛋白酶激活因子1（apoptotic peptidase activating factor 1, APAF-1）的表达并与之结合，促进上皮细胞凋亡，同时诱导APAF-1发生内含子滞留型可变剪接。据我们所知，本研究首次报道了M. bovis通过劫持RBMX2，促进下游APAF-1的内含子剪接，进而调控宿主上皮细胞的凋亡过程。上述研究成果可为新型结核病防控策略的开发提供重要理论参考。