Short-chain fatty acids activate the glutamate-dependent acid resistance system via the tRNA-modifying Mnm pathway in Escherichia coli

Acid resistance is an important trait of enterobacteria, enabling survival under extreme gastric acidity (pH < 2.5) and the moderately acidic conditions in the large intestine (pH 5-6). In Escherichia coli, the glutamate-dependent acid resistance (Gad) system-comprising the glutamate decarboxylases GadA and GadB, and the antiporter GadC-is the major defense system against acid stress. Regulation of the Gad system is highly complex and involves multiple regulatory factors. In this study, we investigated the dynamics of gadA/gadB expression and GadA/GadB protein synthesis in E. coli MG1655 using a controlled bioreactor cultivation. Our results reveal that acid stress, oxygen limitation, and the stationary phase are the stimuli for Gad activation. Notably, acetic acid and other short-chain fatty acids (SCFAs) are superior stimuli of the Gad system than hydrochloric acid. Furthermore, we demonstrate that the tRNA-modifying MnmG/MnmE pathway responds to these SCFAs and stimulates Gad activation, likely via the transcriptional regulator GadW. These findings highlight a previously unrecognized link between SCFAs, tRNA modification, and acid stress resistance.IMPORTANCESurvival of enteric bacteria in the gastrointestinal tract depends on their ability to sense and respond to diverse forms of acid stress. This study identifies oxygen limitation, growth phase, and-most importantly-short-chain fatty acids (SCFAs) as key physiological triggers of the major acid resistance system in Escherichia coli. In addition, we uncover a previously unrecognized connection between SCFAs-sensing, tRNA modification via the MnmG/MnmE pathway, and activation of acid resistance, providing new insight into how metabolic and environmental signals are integrated to promote bacterial survival.