Identification of a MarR subfamily that regulates arsenic resistance genes

In this study, comprehensive analyses were performed to determine the function of an atypical MarR homolog in Achromobacter sp. As-55. Genomic analyses of Achromobacter sp. As-55 showed that this marR is located adjacent to an arsV gene. ArsV is a flavin-dependent monooxygenase that confers resistance to the antibiotic methylarsenite (MAs(III)), the organoarsenic compound roxarsone(III) (Rox(III)), and the inorganic antimonite (Sb(III)). Similar marR genes are widely distributed in arsenic-resistant bacteria. Phylogenetic analyses showed that these MarRs are found in operons predicted to be involved in resistance to inorganic and organic arsenic species, so the subfamily was named MarR ars . MarR ars orthologs have three conserved cysteine residues, which are Cys36, Cys37 and Cys157 in Achromobacter sp. As-55, mutation of which compromises the response to MAs(III)/Sb(III). GFP-fluorescent biosensor assays show that AdMarR ars (MarR protein of Achromobacter deleyi As-55) responds to trivalent As(III) and Sb(III) but not to pentavalent As(V) or Sb(V). The results of RT-qPCR assays show that arsV is expressed constitutively in a marR deletion mutant, indicating that marR represses transcription of arsV . Moreover, electrophoretic mobility shift assays (EMSA) demonstrate that AdMarR ars binds to the promoters of both marR and arsV in the absence of ligands and that DNA binding is relieved upon binding of As(III) and Sb(III). Our results demonstrate that AdMarR ars is a novel As(III)/Sb(III)-responsive transcriptional repressor that controls expression of arsV, which confers resistance to MAs(III), Rox(III) and Sb(III). AdMarR ars and its orthologs form a subfamily of MarR proteins that regulate genes conferring resistance to arsenic-containing antibiotics. IMPORTANCE In this study, a MarR family member, AdMarR ars was shown to regulate the arsV gene, which confers resistance to arsenic-containing antibiotics. It is a founding member of a distinct subfamily that we refer to as MarR ars , regulating genes conferring resistance to arsenic and antimony antibiotic compounds. AdMarR ars was shown to be a repressor containing conserved cysteine residues that are required to bind As(III) and Sb(III), leading to a conformational change and subsequent derepression. Here we show that members of the MarR family are involved in regulating arsenic-containing compounds.

Identification of a MarR subfamily that regulates arsenic resistance genes

In this study, comprehensive analyses were performed to determine the function of an atypical MarR homolog. Genomic analyses showed that this marR is located in an arsenic gene island in Achromobacter sp. As-55 adjacent to an arsV gene. ArsV is a flavin-dependent monooxygenase that confers resistance to the antibiotic methylarsenite (MAs(III)), the organoarsenic compound roxarsone(III) (Rox(III)), and the inorganic antimonite (Sb(III)). Similar marR genes are widely distributed in arsenic-resistant bacteria. Phylogenetic analyses showed that these MarRs are found in operons predicted to be involved in resistance to inorganic and organic arsenic species, so the subfamily was named MarRars. MarRars orthologs have three conserved cysteine residues, which are Cys36, Cys37 and Cys157 in Achromobacter sp. As-55, mutation of which compromises the response to MAs(III)/Sb(III). GFP-fluorescent biosensor assays show that AdMarRars (MarR protein of Achromobacter deleyi As-55) responds to trivalent As(III) and Sb(III) but not to pentavalent As(V) or Sb(V). The results of RT-qPCR assays show that arsV is expressed constitutively in a marR deletion mutant, indicating that marR represses transcription of arsV. Moreover, electrophoretic mobility shift assays (EMSA) demonstrate that AdMarRars binds to the promoters of both marR and arsV in the absence of ligands and that DNA binding is relieved upon binding of As(III) and Sb(III). Our results demonstrate that AdMarRars is a novel As(III)/Sb(III)-responsive transcriptional repressor that controls expression of arsV, which confers resistance to MAs(III), Roxarsone(III) and Sb(III). AdMarRars and its orthologs form a subfamily of MarR proteins that regulate genes conferring resistance to arsenic-containing antibiotics.

1. Isolation and Biochemical Characterization of Arsenic resistant Bacteria from Textile Effluent Contaminated Soil of Jaipur, Rajasthan

Arsenic occurs naturally in the environment, but its uncontrolled liberation from industrial effluents has been imposing adverse effects on the environment. The continuous exposure of the soil is a matter of concern in this study. The soil consortium contains bacterial colonies that resist and adapt the metal toxicants and can in turn help in the bioremediation of such metals from the soil. This study stresses the isolation of arsenic resistant bacteria from the arsenic-contaminated soil. The Soil Sample was collected from the Sanganer area of Jaipur, Rajasthan. The soil in this area received the textile discharge from the industries located nearby. Four arsenic resistant bacteria were isolated from the sample which showed a high tolerance level towards arsenic and was able to grow in the presence of arsenic in-vitro. The Minimum inhibitory concentration was also determined for the strains against arsenite. The highest MIC was found to be 16mM of arsenite, which concludes tolerable limits of the strains. The Biochemical and Morphological characterization of the isolates was also conducted. The Four isolates also showed resistance towards various other metals like cadmium, cobalt, lead, zinc, mercury, chromium, and tin. The isolates on biochemical characterization were found to belong to the following Genus: Moraxella, Azomonas, Acetobactor and Corynebacterium. This resistance capacity of the isolates depicts their potential to bioremediate the toxicity of the arsenic in the environment.