Department of Microbiology, University of Washington, Seattle, WA, USA.
Interdisciplinary Program of Pathobiology, Department of Global Health, University of Washington, Seattle, WA, USA.
Nat Microbiol. 2021 Jan;6(1):44-50. doi: 10.1038/s41564-020-00810-x. Epub 2020 Nov 16.
Transposon-based strategies provide a powerful and unbiased way to study the bacterial stress response, but these approaches cannot fully capture the complexities of network-based behaviour. Here, we present a network-based genetic screening approach: the transcriptional regulator-induced phenotype (TRIP) screen, which we used to identify previously uncharacterized network adaptations of Mycobacterium tuberculosis to the first-line anti-tuberculosis drug isoniazid (INH). We found regulators that alter INH susceptibility when induced, several of which could not be identified by standard gene disruption approaches. We then focused on a specific regulator, mce3R, which potentiated INH activity when induced. We compared mce3R-regulated genes with baseline INH transcriptional responses and implicated the gene ctpD (Rv1469) as a putative INH effector. Evaluating a ctpD disruption mutant demonstrated a previously unknown role for this gene in INH susceptibility. Integrating TRIP screening with network information can uncover sophisticated molecular response programs.
转座子策略为研究细菌应激反应提供了一种强大且无偏倚的方法，但这些方法无法完全捕捉到基于网络的行为的复杂性。在这里，我们提出了一种基于网络的遗传筛选方法：转录调节因子诱导表型（TRIP）筛选，我们用它来鉴定结核分枝杆菌对一线抗结核药物异烟肼（INH）的以前未表征的网络适应。我们发现了一些诱导时会改变 INH 敏感性的调节剂，其中一些调节剂无法通过标准基因敲除方法来识别。然后，我们将重点放在一个特定的调节剂 mce3R 上，它在诱导时增强了 INH 的活性。我们将 mce3R 调节的基因与 INH 的基础转录反应进行了比较，并将基因 ctpD（Rv1469）作为潜在的 INH 效应物。评估 ctpD 缺失突变体表明该基因在 INH 敏感性中具有以前未知的作用。将 TRIP 筛选与网络信息相结合可以揭示复杂的分子反应程序。
