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Transcriptional Regulator AcrR Increases Ethanol Tolerance through Regulation of Fatty Acid Synthesis in Lactobacillus plantarum.

ABSTRACT: Lactobacillus plantarum is a versatile bacterium with significant adaptability to harsh habitats containing excessive ethanol concentrations. It was found that the L. plantarum NF92-TetR/AcrR family regulator, AcrR, significantly enhanced the growth rate of this lactic acid bacterium in the presence of ethanol. Through screening 172 ethanol-resistant related genes by electrophoretic mobility shift and quantitative reverse transcription-PCR (RT-qPCR) assays, six genes were identified to be regulated by AcrR under ethanol stress. Among these was a gene coding for a 3-hydroxyacyl-ACP dehydratase (fabZ1) regulated by AcrR under ethanol stress. AcrR regulated fabZ1 under ethanol stress by binding to its promoter, P _fabZ1 DNase I footprinting analysis indicated that there were two specific AcrR binding sites on P _fabZ1 RT-PCR results showed fabZ1 could cotranscribe with its downstream 12 genes and conform a fatty acid de novo biosynthesis (fab) gene cluster under the control of P _fabZ1 Both RT-qPCR of the fab gene cluster in acrR knockout and overexpression strains and fatty acid methyl ester analysis of the acrR knockout strain showed that AcrR could promote fatty acid synthesis in L. plantarum NF92. Membrane fluorescence anisotropy analysis of acrR knockout and overexpression strains showed that AcrR could increase membrane fluidity under ethanol stress. Thus, AcrR could regulate fatty acid synthesis and membrane fluidity to promote the adaption of L. plantarum NF92 to a high ethanol concentration.IMPORTANCE Ethanol tolerance is essential for L. plantarum strains living in substances with more than 9% ethanol, such as wine and beer. The details regarding how L. plantarum adapts to ethanol are still lacking. This study demonstrates that AcrR regulates the de novo synthesis of fatty acids in L. plantarum adapting to toxic levels of ethanol. We also identified the ability of the TetR/AcrR family regulator to bind to the fatty acid biosynthesis gene promoter, P _fabZ1 , in L. plantarum and defined the binding sites. This finding facilitates the induction of the adaptation of L. plantarum strains to ethanol for food fermentation applications.

SUBMITTER: Yang X

PROVIDER: S-EPMC6821977 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Lactobacillus plantarum is a versatile bacterium that occupies a wide range of environmental niches. In this study, we found that a bifunctional aldehyde-alcohol dehydrogenase-encoding gene, adhE, was responsible for L. plantarum being able to utilize mannitol and sorbitol through cross-regulation by two DNA-binding regulators. In L. plantarum NF92, adhE was greatly induced, and the growth of an adhE-disrupted (ΔadhE) strain was repressed when sorbitol or mannitol instead of glucose was used as a carbon source. The results of enzyme activity and metabolite assays demonstrated that AdhE could catalyze the synthesis of ethanol in L. plantarum NF92 when sorbitol or mannitol was used as the carbon source. AcrR and Rex were two transcriptional factors screened by an affinity isolation method and verified to regulate the expression of adhE DNase I footprinting assay results showed that they shared a binding site (GTTCATTAATGAAC) in the adhE promoter. Overexpression and knockout of AcrR showed that AcrR was a novel regulator to promote the transcription of adhE The activator AcrR and repressor Rex may cross-regulate adhE when L. plantarum NF92 utilizes sorbitol or mannitol. Thus, a model of the control of adhE by AcrR and Rex during L. plantarum NF92 utilization of mannitol or sorbitol was proposed.IMPORTANCE The function and regulation of AdhE in the important probiotic genus Lactobacillus are rarely reported. Here we demonstrated that AdhE is responsible for sorbitol and mannitol utilization and is cross-regulated by two transcriptional regulators in L. plantarum NF92, which had not been reported previously. This is important for L. plantarum to compete and survive in some harsh environments in which sorbitol or mannitol could be used as carbon source. A novel transcriptional regulator AcrR was identified to be important to promote the expression of adhE, which was unknown before. The cross-regulation of adhE by AcrR and Rex is important to balance the level of NADH in the cell during sorbitol or mannitol utilization.

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Transcriptional Regulator AcrR Increases Ethanol Tolerance through Regulation of Fatty Acid Synthesis in Lactobacillus plantarum.

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