Aromatic ring-hydroxylating dioxygenases (ARHDs) play a crucial role in the aerobic biodegradation of both natural and anthropogenic aromatic compounds. Although their ability to process contaminants is not entirely understood, it is thought to have evolved from the transformation of structurally similar secondary plant metabolites (SPMs). Hence, to investigate this connection, we tested a variety of SPMs from the monoterpene and flavonoid classes as carbon sources and transcriptional effectors of several phylogenetically distant ARHD genes involved in the degradation of aromatic pollutants. Specifically, we focused on bphA1, nahA1 and phtA1 in Rhodococcus opacus C1, whose genomic analysis is also presented hereinafter, and bphA1a, nahA1-bphA1b and etbA1ab in Rhodococcus sp. WAY2. Whilst induction was only observed with (R)-carvone for bphA1a and nahA1-bphA1b of strain WAY2, and with p-cymene for nahA1 and nahA1-bphA1b of strains C1 and WAY2, respectively, an extensive inhibition by flavonoids was observed for most of the genes in both strains. To the best of our knowledge, our study is the first to report the effect of flavonoids and monoterpenes on the transcription of nahA1, etbA1 and phtA1 genes. In addition, we show that, in contrast to pseudomonads, many flavonoids inhibit the transcription of the ARHD genes in rhodococci. Thus, our work provides a new perspective on flavonoids as the transcriptional effectors of ARHDs, highlighting the significant variability of these enzymes and the divergent responses that they elicit. Moreover, our results contribute to understanding the complex interactions between microorganisms and SPMs and provide insights into the molecular basis of a number of them.

• Klíčová slova
• Rhodococcus, aromatic pollutants, aromatic ring-hydroxylating dioxygenases, biodegradation, flavonoids, monoterpenes,
• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• biodegradace MeSH
• dioxygenasy * genetika   metabolismus   MeSH
• flavonoidy * farmakologie   metabolismus   MeSH
• fylogeneze MeSH
• genetická transkripce účinky léků   MeSH
• monoterpeny * farmakologie   metabolismus   MeSH
• regulace genové exprese u bakterií účinky léků   MeSH
• Rhodococcus * genetika   účinky léků   enzymologie   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH
• dioxygenasy * MeSH
• flavonoidy * MeSH
• monoterpeny * MeSH

In this study, the diversity of bphA genes was assessed in a 13C-enriched metagenome upon stable isotope probing (SIP) of microbial populations in legacy PCB-contaminated soil with 13C-biphenyl (BP). In total, 13 bphA sequence variants (SVs) were identified in the final amplicon dataset. Of these, one SV comprised 59% of all sequences, and when it was translated into a protein sequence, it exhibited 87, 77.4, and 76.7% identity to its homologs from Pseudomonas furukawaii KF707, Cupriavidus sp. WS, and Pseudomonas alcaliphila B-367, respectively. This same BphA sequence also contained unusual amino acid residues, Alanine, Valine, and Serine in region III, which had been reported to be crucial for the substrate specificity of the corresponding biphenyl dioxygenase (BPDO), and was accordingly designated BphA_AVS. The DNA locus of 18 kbp containing the BphA_AVS-coding sequence retrieved from the metagenome was comprised of 16 ORFs and was most likely borne by Paraburkholderia sp. The BPDO corresponding to bphAE_AVS was cloned and heterologously expressed in E. coli, and its substrate specificity toward PCBs and a spectrum of flavonoids was assessed. Although depleting a rather narrow spectrum of PCB congeners, the efficient transformation of flavone and flavanone was demonstrated through dihydroxylation of the B-ring of the molecules. The homology-based functional assignment of the putative proteins encoded by the rest of ORFs in the AVS region suggests their potential involvement in the transformation of aromatic compounds, such as flavonoids. In conclusion, this study contributes to the body of information on the involvement of soil-borne BPDOs in the metabolism of flavonoid compounds, and our paper provides a more advanced context for understanding the interactions between plants, microbes and anthropogenic compounds in the soil.

• Klíčová slova
• aromatic ring-hydroxylating dioxygenases, biphenyl dioxygenase, flavonoids, functional metagenomics, polychlorinated biphenyls, secondary plant metabolites,
• Publikační typ
• časopisecké články MeSH

The involvement of bacterial aromatic ring-hydroxylating dioxygenases (ARHDs) in the degradation of aromatic pollutants, such as polychlorinated biphenyls (PCBs), has been well studied. However, there is considerable speculation as to the origin of this ability. One hypothesis is centered on a connection between the ability to degrade aromatic pollutants and the necessity of soil bacteria to cope with and/or utilize secondary plant metabolites (SPMs). To investigate this connection, we researched the involvement of biphenyl 2,3-dioxygenase (BPDO), an ARHD essential for the degradation of PCBs, in the metabolism of SPMs in the soil bacterium Pseudomonas alcaliphila JAB1, a versatile degrader of PCBs. We demonstrated the ability of the strain JAB1 to transform a variety of SPMs, namely the flavonoids apigenin, flavone, flavanone, naringenin, fisetin, quercetin, morin, and catechin, caffeic acid, trans-cinnamic acid, and the monoterpenes (S)-limonene and (R)-carvone. Of those, the transformation of flavone, flavanone, and (S)-limonene was conditioned by the activity of JAB1-borne BPDO and thus was researched in more detail, and we found evidence for the limonene monooxygenase activity of the BPDO. Furthermore, the bphA gene in the strain JAB1 was demonstrated to be induced by a wide range of SPMs, with monoterpenes being the strongest inducers of the SPMs tested. Thus, our findings contribute to the growing body of evidence that ARHDs not only play a role in the catabolism of aromatic pollutants, but also of natural plant-derived aromatics, and this study supports the hypothesis that ARHDs participate in ecological processes mediated by SPMs.

• Klíčová slova
• aromatic ring-hydroxylating dioxygenases, biphenyl dioxygenase, monoterpenes, phenolics, secondary plant metabolites,
• Publikační typ
• časopisecké články MeSH

Certain industrial chemicals accumulate in the environment due to their recalcitrant properties. Bioremediation uses the capability of some environmental bacteria to break down these chemicals and attenuate the pollution. One such bacterial strain, designated Pvy, was isolated from sediment samples from a lagoon in Romania located near an oil refinery due to its capacity to degrade dibenzofuran (DF). The genome sequence of the Pvy strain was obtained using an Oxford Nanopore MiniION platform. According to the consensus 16S rRNA gene sequence that was compiled from six 16S rRNA gene copies contained in the genome and orthologous average nucleotide identity (OrthoANI) calculation, the Pvy strain was identified as Pseudomonas veronii, which confirmed the identification obtained with the aid of MALDI-TOF mass spectrometry and MALDI BioTyper. The genome was analyzed with respect to enzymes responsible for the overall biodegradative versatility of the strain. The Pvy strain was able to derive carbon from naphthalene (NP) and several aromatic compounds of natural origin, including salicylic, protocatechuic, p-hydroxybenzoic, trans-cinnamic, vanillic, and indoleacetic acids or vanillin, and was shown to degrade but not utilize DF. In total seven loci were found in the Pvy genome, which enables the strain to participate in the degradation of these aromatic compounds. Our experimental data also indicate that the transcription of the NP-dioxygenase α-subunit gene (ndoB), carried by the plasmid of the Pvy strain, is inducible by DF. These features make the Pvy strain a potential candidate for various bioremediation applications.

• Klíčová slova
• Pseudomonas veronii strain Pvy, biodegradation, denitrification, dibenzofuran, dioxygenase, heavy-metal tolerance, nanopore technology, organic phosphate mineralization, whole-genome sequencing,
• MeSH
• biodegradace MeSH
• dibenzofurany * MeSH
• genomika * MeSH
• Pseudomonas MeSH
• RNA ribozomální 16S MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• dibenzofurany * MeSH
• RNA ribozomální 16S MeSH