A novel RofA-family transcriptional regulator, GadR, controls the development of acid resistance inListeria monocytogenes

Abstract

AbstractStomach acid provides a significant innate barrier to the entry of the food-borne pathogenListeria monocytogenesinto the human gastrointestinal tract. A key determinant of acid resistance in this bacterium is the conserved glutamate decarboxylase system, GadD2 (encoded by thegadT2D2operon), which helps to maintain the intracellular pH during exposure to gastric acid. In this study, we identified a premature stop codon in a gene located immediately downstream of thegadT2D2operon that was highly linked to an acid sensitive phenotype. When this open reading frame was restored through homologous recombination an acid resistant phenotype resulted. Through a series of genetic, transcriptomic and survival experiments we established that this gene, which we designatedgadR, encodes a transcriptional regulator of thegadT2D2operon. GadR belongs to the RofA family of regulators, primarily found in the streptococci, where they are involved in regulating virulence. The data further showed thatgadRplays a critical role in the development of acid resistance in response to mild acid exposure, a response that is known as the adaptive acid tolerance response (ATR). A deletion analysis of thegadT2D2promoter region identified two 18bp palindromic sequences that are required for the GadR-mediated induction ofgadT2D2, suggesting that they act as binding sites for GadR. Overall, this study uncovers a new RofA-like regulator of acid resistance inL. monocytogenesthat plays a significant role in both growth phase-dependent and ATR mediated acid resistance and accounts for previously observed strain-to-strain differences in survival at low pH. The findings have important implications for understanding the behavior ofL. monocytogenesin acidic environments and identify a potential target for improved control of this important pathogen.Author summaryThe ability to survive the acidic conditions found in the stomach is a key trait enabling the food-borne pathogenListeria monocytogenesto gain access to mammalian gastrointestinal tract, where it can initiate an infection. Little is currently known about how acid resistance is regulated in this pathogen and why this trait is highly variable between strains. Here we used genomic sequences from a collection ofL. monocytogenesstrains with known differences in acid survival to identify a novel transcriptional regulator controlling acid resistance, which we call GadR. The regulator belongs to a family of regulators previously found only in a small group of bacterial pathogens including the streptococci, where they are involved in regulating virulence properties. We show that GadR is the dominant regulator of acid resistance inL. monocytogenesand that variability in its gene sequence accounts for previously observed differences between strains in this trait. Together these findings significantly advance our understanding of how this important pathogen copes with acid stress and suggests a potential molecular target to better control it in the human food-chain.