Transcriptional Regulation of the Peripheral Pathway for the Anaerobic Catabolism of Toluene and m-Xylene in Azoarcus sp. CIB.

Blas Blázquez,Eduardo Díaz,Manuel Carmona

doi:10.3389/fmicb.2018.00506

Blas Blázquez, Eduardo Díaz + Show 1 more

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https://doi.org/10.3389/fmicb.2018.00506

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Abstract

Alkylbenzenes, such as toluene and m-xylene, are an important class of contaminant hydrocarbons that are widespread and tend to accumulate in subsurface anoxic environments. The peripheral pathway for the anaerobic oxidation of toluene in bacteria consists of an initial activation catalyzed by a benzylsuccinate synthase (encoded by bss genes), and a subsequent modified β-oxidation of benzylsuccinate to benzoyl-CoA and succinyl-CoA (encoded by bbs genes). We have shown here that the bss and bbs genes, which are located within an integrative and conjugative element, are essential for anaerobic degradation of toluene but also for m-xylene oxidation in the denitrifying beta-proteobacterium Azoarcus sp. CIB. New insights into the transcriptional organization and regulation of a complete gene cluster for anaerobic catabolism of toluene/m-xylene in a single bacterial strain are presented. The bss and bbs genes are transcriptionally coupled into two large convergent catabolic operons driven by the PbssD and PbbsA promoters, respectively, whose expression is inducible when cells grow anaerobically in toluene or m-xylene. An adjacent tdiSR operon driven by the PtdiS promoter encodes a putative two-component regulatory system. TdiR behaves as a transcriptional activator of the PbssD, PbbsA, and PtdiS promoters, being benzylsuccinate/(3-methyl)benzylsuccinate, rather than toluene/m-xylene, the inducers that may trigger the TdiS-mediated activation of TdiR. In addition to the TdiSR-based specific control, the expression of the bss and bbs genes in Azoarcus sp. CIB is under an overimposed regulation that depends on certain environmental factors, such as the presence/absence of oxygen or the availability of preferred carbon sources (catabolite repression). This work paves the way for future strategies toward the reliable assessment of microbial activity in toluene/m-xylene contaminated environments.

Highlights

Aromatic compounds such as alkylbenzenes, e.g., toluene, xylenes, ethylbenzene and benzene, are an important class of contaminants that are prominently placed amongst the US Agency for Toxic Substances and Disease Registry’s list of priority pollutants because of their carcinogenic and/or neurotoxic effects to humans
Since the toluene peripheral pathway was shown to be inducible when bacteria grow in the presence of toluene (Coschigano, 2000; Achong et al, 2001; Hermuth et al, 2002; Kube et al, 2004; Kühner et al, 2005), we checked whether the expression of the bss-bbs genes was inducible when Azoarcus sp
The analysis of the RT-PCR amplification products revealed that the expression of the bssA and bbsA genes is induced when the cells grow in the presence of toluene or m-xylene as sole carbon sources (Figures 3A,B), suggesting their participation in the anaerobic degradation of both aromatic hydrocarbons

Summary

Introduction

Aromatic compounds such as alkylbenzenes, e.g., toluene, xylenes, ethylbenzene and benzene, are an important class of contaminants that are prominently placed amongst the US Agency for Toxic Substances and Disease Registry’s list of priority pollutants because of their carcinogenic and/or neurotoxic effects to humans. Several isolates capable of anaerobic toluene degradation have been described since including both facultative anaerobes, e.g., beta-proteobacteria of the Azoarcus, “Aromatoleum,” Thauera, Georgfuchsia, Herminiimonas genera, and alpha-proteobacteria of the Magnetospirillum genus, and obligate anaerobes, e.g., delta-proteobacteria of the Geobacter, Desulfobacula and Desulfobacterium genera, and some clostridial strains (Wöhlbrand et al, 2013; Bozinovski et al, 2014; Kim et al, 2014; Strijkstra et al, 2014; Rabus et al, 2016; Lueders, 2017; Meyer-Cifuentes et al, 2017). Most of the studies on anaerobic toluene degradation have been made on the denitrifying bacteria Thauera aromatica K172 and T1 strains, Azoarcus sp. strain T and “Aromatoleum aromaticum” EbN1 strain (Coschigano and Young, 1997; Leuthner et al, 1998; Krieger et al, 1999; Achong et al, 2001; Hermuth et al, 2002; Leutwein and Heider, 2002; Chakraborty and Coates, 2004; Coschigano and Bishop, 2004; Kube et al, 2004; Kühner et al, 2005)

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