Global transcriptional response of Methylorubrum extorquens to formaldehyde stress includes both overlapping and unique gene sets in comparison to antibiotic translational inhibition and expands the role of EfgA

The potency and indiscriminate nature of formaldehyde reactivity upon biological molecules make it a universal stressor. However, some organisms such as Methylorubrum extorquens possess means to rapidly and effectively mitigate formaldehyde-induced damage. EfgA is a recently identified formaldehyde sensor predicted to halt translation in response to elevated formaldehyde as a means to protect cells. Herein, we investigate growth, formaldehyde consumption, and changes in gene expression to better understand how M. extorquens responds to formaldehyde with and without the EfgA-formaldehyde-mediated translational response, and how this mechanism compares to other forms of translation inhibition. These distinct mechanisms of translation inhibition have notable differences: they each involve different specific players and in addition, formaldehyde also acts as a general, multi-target stressor and a potential carbon source. Here, we present findings demonstrating that in addition to its characterized impact on translation, functional EfgA also allows for a rapid and robust transcriptional response to formaldehyde and that removal of efgA leads to heightened proteotoxic and genotoxic stress in the presence of increased formaldehyde levels. As our previous work suggested that formaldehyde is proteotoxic in M. extorquens, we accurately predicted that strains lacking efgA would experience increased protein damage. We also found that many downstream consequences of translation inhibition were shared by EfgA-formaldehyde- and kanamycin-mediated translation inhibition. Our work to modularize the transcriptional response uncovered additional layers of regulatory control enacted by functional EfgA upon experiencing formaldehyde stress, and further demonstrate the importance this protein plays at both transcriptional and translational levels in this model methylotroph. Time courses of gene expression after treatment with two different translational inhibitors (kanamycin and formaldehyde). The responses of wild type and an ΔefgA mutant of Methylorubrum extorquens were compared.

甲醛对生物分子的反应活性极强且无选择性，使其成为一种通用胁迫因子。然而，部分生物如扭脱甲基杆菌（Methylorubrum extorquens）具备快速且有效缓解甲醛诱导损伤的能力。EfgA是新近发现的甲醛感受器，据预测可响应甲醛浓度升高而阻断翻译过程，以此保护细胞。
本研究通过探究生长情况、甲醛消耗水平以及基因表达变化，以期更深入理解扭脱甲基杆菌在存在或缺失EfgA介导的甲醛翻译阻断通路时对甲醛的响应机制，并对比该机制与其他翻译抑制方式的差异。此类不同的翻译抑制机制存在显著区别：它们各自涉及不同的特异性调控因子，此外，甲醛同时也是一种多靶点通用胁迫因子与潜在碳源。
本研究结果显示，除已被证实的对翻译过程的影响外，功能性EfgA还可介导对甲醛的快速且强效的转录响应；而敲除efgA会在甲醛浓度升高时加剧细胞的蛋白毒性与遗传毒性胁迫。正如我们此前研究表明甲醛对扭脱甲基杆菌具有蛋白毒性，我们精准预测到缺失efgA的菌株会遭受更严重的蛋白质损伤。
我们还发现，翻译抑制的诸多下游效应在EfgA-甲醛介导与卡那霉素（kanamycin）介导的翻译抑制过程中具有共通性。本研究通过模块化解析转录响应，揭示了功能性EfgA在遭遇甲醛胁迫时所介导的额外层级的调控机制，并进一步证实了该蛋白在这一模式甲基营养菌中于转录与翻译层面均发挥关键作用。
本研究对比了野生型与ΔefgA缺失突变株扭脱甲基杆菌经两种不同翻译抑制剂（卡那霉素与甲醛）处理后的基因表达时间进程。