Transcriptomics reveal strain-specific metabolic strategies for acid resistance and gamma-aminobutyric acid (GABA) production in Levilactobacillus brevis.

Background: Of the many neurotransmitters in humans, gamma-aminobutyric acid (GABA) shows potential for improving several mental health indications such as stress and anxiety. The microbiota-gut-brain axis is an important pathway for GABAergic effects, as microbially-secreted GABA within the gut can affect host mental functionhealth outcomes. Understanding the molecular characteristics of GABA production by microbes within the gut can offer insight to novel therapies for mental health. Results: Three strains of Levilactobacillus brevis with syntenous glutamate decarboxylase (GAD) operons were evaluated for overall growth, glutamate utilization, and GABA production in typical synthetic growth media supplemented with monosodium glutamate (MSG). Levilactobacillus brevis Lbr-6108 (Lbr-6108) and Levilactobacillus brevis Lbr-35 (Lbr-35) had similar growth profiles but differed significantly in GABA secretion and acid resistance. Lbr-6108 produced GABA early, within the growth phase, and produced significantly more GABA than Lbr-35 and the type strain Levilactobacillus brevis ATCC 14689 after the stationary phase. The global gene expression during GABA production was determined by RNA sequencing at several timepoints. The GAD operon, responsible for GABA production and secretion, activated in Lbr-6108 after only six hours of fermentation and continued throughout the stationary phase. Furthermore, Lbr-6108 activated many different acid resistance mechanisms concurrently, which contribute to acid tolerance and energy production. In contrast, Lbr-35, which has a genetically similar GAD operon, including two copies of the GAD gene, showed no upregulation of the GAD operon, even when cultured with MSG. Conclusions: This study is the first to evaluate whole transcriptome changes in L. brevis during GABA production over multiple timepoints. The concurrent expression of multiple acid-resistance mechanisms reveals niche-specific metabolic functionality between common human commensals and highlights the complex regulation of GABA metabolism in this important microbial species. Furthermore, the increased and rapid GABA production of Lbr-6108 highlights the strain’s potential as a therapeutic and the overall value of screening microbes for effector molecule output. Gene expression of Lbr-35 and Lbr-6108 timeseries grown in media with and without additional monosodium glutamate.

背景：在人体众多神经递质中，γ-氨基丁酸（gamma-aminobutyric acid, GABA）在改善应激、焦虑等多种心理健康相关病症方面展现出应用潜力。微生物群-肠-脑轴（microbiota-gut-brain axis）是介导GABA能作用的重要通路，因为肠道内微生物分泌的GABA可影响宿主的心理功能与健康结局。解析肠道微生物合成GABA的分子特征，可为心理健康领域的新型治疗策略提供新思路。
结果：本研究针对3株携带同源性谷氨酸脱羧酶（glutamate decarboxylase, GAD）操纵子的短乳杆菌（Levilactobacillus brevis）展开评估，在添加谷氨酸钠（monosodium glutamate, MSG）的典型合成培养基中，检测其整体生长态势、谷氨酸利用效率与GABA合成能力。短乳杆菌Lbr-6108（Lbr-6108）与短乳杆菌Lbr-35（Lbr-35）的生长概况相似，但在GABA分泌能力与耐酸性方面存在显著差异。Lbr-6108在生长阶段早期即可合成GABA，且在稳定期后产生的GABA量显著高于Lbr-35以及模式菌株短乳杆菌ATCC 14689。本研究通过多个时间点的RNA测序（RNA sequencing）分析了GABA合成过程中的全基因表达谱。负责GABA合成与分泌的GAD操纵子仅在发酵6小时后便在Lbr-6108中被激活，并贯穿整个稳定期持续表达。此外，Lbr-6108可同时激活多种不同的耐酸机制，这些机制有助于提升酸耐受性与能量产生效率。与之形成对比的是，尽管Lbr-35携带遗传相似度较高的GAD操纵子（包含2个GAD基因拷贝），但即使在添加MSG的培养基中培养，其GAD操纵子也未出现表达上调。
结论：本研究首次在多个时间点下评估了短乳杆菌（Levilactobacillus brevis）在GABA合成过程中的全转录组变化。多种耐酸机制的协同表达揭示了常见人类共生菌之间的生态位特异性代谢功能，并凸显了这一重要微生物物种中GABA代谢的复杂调控机制。此外，Lbr-6108所具备的高效且快速的GABA合成能力，凸显了该菌株作为治疗制剂的应用潜力，同时也证明了通过筛选微生物获取效应分子的整体研究价值。本数据集包含在添加与未添加额外谷氨酸钠的培养基中培养的Lbr-35与Lbr-6108的时间序列基因表达数据。