Divergent roles of CprK paralogues from Desulfitobacterium hafniense in activating gene expression

Gábor, Krisztina; Sene, Kagnew Hailesellasse; Smidt, Hauke; Vos, Willem M. de; Oost, John van der

doi:10.1099/mic.0.2008/021584-0

Cited by 20 publications

(26 citation statements)

References 42 publications

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“…This was for the reason that functional screening of rdhA seems to be difficult because functional expression of rdhA genes in E. coli has not been successful [53,54]. On the other hand, the utility of an in vivo transcription assay of the transcriptional regulator CprK in E. coli has been proved [55], indicating that it might be possible to explore genes involved in organohalide respiration using SIGEX transcriptional screening. Contrary to the results of the 16S rRNA gene sequence analysis, which indicated that Desulfuromonadales species predominated in the dehalogenation-positive enrichment cultures, SIGEX suggested the presence of a phylogenetically diverse assortment of clones putatively belong to the Gammaproteobacteria, Deltaproteobacteria, Dehalococcoidetes, Deferribacteres and Methanomicrobia.…”

Section: Discussionmentioning

confidence: 99%

Distribution of dehalogenation activity in subseafloor sediments of the Nankai Trough subduction zone

Futagami

Morono

Terada

et al. 2013

Phil. Trans. R. Soc. B

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Halogenated organic matter buried in marine subsurface sediment may serve as a source of electron acceptors for anaerobic respiration of subseafloor microbes. Detection of a diverse array of reductive dehalogenase-homologous ( rdhA ) genes suggests that subseafloor organohalide-respiring microbial communities may play significant ecological roles in the biogeochemical carbon and halogen cycle in the subseafloor biosphere. We report here the spatial distribution of dehalogenation activity in the Nankai Trough plate-subduction zone of the northwest Pacific off the Kii Peninsula of Japan. Incubation experiments with slurries of sediment collected at various depths and locations showed that degradation of several organohalides tested only occurred in the shallow sedimentary basin, down to 4.7 metres below the seafloor, despite detection of rdhA in the deeper sediments. We studied the phylogenetic diversity of the metabolically active microbes in positive enrichment cultures by extracting RNA, and found that Desulfuromonadales bacteria predominate. In addition, for the isolation of genes involved in the dehalogenation reaction, we performed a substrate-induced gene expression screening on DNA extracted from the enrichment cultures. Diverse DNA fragments were obtained and some of them showed best BLAST hit to known organohalide respirers such as Dehalococcoides , whereas no functionally known dehalogenation-related genes such as rdhA were found, indicating the need to improve the molecular approach to assess functional genes for organohalide respiration.

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Section: Discussionmentioning

confidence: 99%

Distribution of dehalogenation activity in subseafloor sediments of the Nankai Trough subduction zone

Futagami

Morono

Terada

et al. 2013

Phil. Trans. R. Soc. B

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“…Most of rdh gene clusters are also associated with one rdhK subunit in various orientations. The rdhK-encoded proteins clearly belong to the large family of CRP/Fnr regulatory proteins from which CprK members of Desulfitobacterium dehalogenans and Desulfitobacterium hafniense DCB-2 were extensively studied and represent the paradigmatic DNA-binding regulatory protein for the respective chlorophenol reductive dehalogenase (cpr) operons [48][49][50][51][52][53][54][55][56] figure 3). First, a strong correlation could be established between the level of sequence identity (see also electronic supplementary material, table S7) and the genetic organization of the predicted rdh operons.…”

Section: (I) Multiple Reductive Dehalogenase Homologue Gene Clusters mentioning

confidence: 99%

The restricted metabolism of the obligate organohalide respiring bacterium Dehalobacter restrictus: lessons from tiered functional genomics

Rupakula

Kruse

Boeren

et al. 2013

Phil. Trans. R. Soc. B

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Dehalobacter restrictus strain PER-K23 is an obligate organohalide respiring bacterium, which displays extremely narrow metabolic capabilities. It grows only via coupling energy conservation to anaerobic respiration of tetra- and trichloroethene with hydrogen as sole electron donor. Dehalobacter restrictus represents the paradigmatic member of the genus Dehalobacter , which in recent years has turned out to be a major player in the bioremediation of an increasing number of organohalides, both in situ and in laboratory studies. The recent elucidation of the D. restrictus genome revealed a rather elaborate genome with predicted pathways that were not suspected from its restricted metabolism, such as a complete corrinoid biosynthetic pathway, the Wood–Ljungdahl (WL) pathway for CO 2 fixation, abundant transcriptional regulators and several types of hydrogenases. However, one important feature of the genome is the presence of 25 reductive dehalogenase genes, from which so far only one, pceA , has been characterized on genetic and biochemical levels. This study describes a multi-level functional genomics approach on D. restrictus across three different growth phases. A global proteomic analysis allowed consideration of general metabolic pathways relevant to organohalide respiration, whereas the dedicated genomic and transcriptomic analysis focused on the diversity, composition and expression of genes associated with reductive dehalogenases.

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“…PCP was then added at 30 M, whereas 2,4,6-TCP was added at 0. 125,5, 25, and 50 M and 3,3,6, 30, and 60 M (final concentration). The cultures were incubated at 30°C for 12 h. For the temperature experiment, the bottles were incubated for 3 h at 22, 30, or 37°C prior to addition of 3,5-DCP at 60 M. The bottles were further incubated for 12 h at these temperatures.…”

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confidence: 99%

Quantitative Analysis of the Relative Transcript Levels of Four Chlorophenol Reductive Dehalogenase Genes in Desulfitobacterium hafniense PCP-1 Exposed to Chlorophenols

Bisaillon

Beaudet

Lépine

et al. 2011

Appl Environ Microbiol

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Relative to those of unexposed cultures, the transcript levels of the four CprA-type reductive dehalogenase genes (cprA2, cprA3, cprA4, and cprA5) in Desulfitobacterium hafniense PCP-1 were measured in cultures exposed to chlorophenols. In 2,4,6-trichlorophenol-amended cultures, cprA2 and cprA3 were upregulated, as was cprA5, but concomitantly with the appearance of 2,4-dichlorophenol (DCP). In 3,5-DCP-amended cultures, only cprA5 was upregulated. In pentachlorophenol-amended cultures grown for 12 h, cprA2 and cprA3 were upregulated but not cprA5. cprA4 was not upregulated significantly in cultures containing any tested chlorophenols.Desulfitobacterium hafniense PCP-1 is able to dehalogenate pentachlorophenol (PCP) and several chlorophenols (3) by hydrogenolytic reductive dehalogenation (5,9,15,16,19). This dehalogenation involves two different enzymatic systems: the ortho-dechlorinating system and the meta-/para-dechlorinating system (2). At least four open reading frames (cprA2, cprA3, cprA4, and cprA5) encoding CprA-type reductive dehalogenases (RDases) were identified in this strain (8). We previously showed that cprA3 in strain PCP-1 encodes an RDase with ortho-dechlorinating activity toward highly chlorinated phenols such as PCP, tetrachlorophenols, and several trichlorophenols (TCPs) (1). We also determined that cprA5 encodes an RDase with meta-and para-dechlorinating activities toward several TCPs and dichlorophenols (DCPs) (17). The functions of the putative RDases encoded by the cprA2 and cprA4 genes have yet to be determined. We hypothesized that 3,5-DCP (a meta-/ para-dechlorinating activity inducer) and 2,4,6-TCP (an orthodechlorinating activity inducer) modulate differently the level of transcription of these genes in strain PCP-1. In this study, we examined by real-time reverse transcription quantitative PCR (RT-qPCR) changes in the transcript levels of cprA2 to cprA5 following the addition of inducer 2,4,6-TCP or 3,5-DCP to D. hafniense PCP-1 cultures and their response to PCP exposure.Culture conditions. D. hafniense PCP-1 (ATCC 700357) was cultivated anaerobically in either 70-ml serum vials or 1-liter bottles containing, respectively, 35 or 500 ml of mineral salt medium with pyruvate and yeast extract as described previously (8). In all experiments, the medium was inoculated with 5% (vol/vol) of an exponentially growing culture. For 2,4,6-TCP and 3,5-DCP exposure experiments, strain PCP-1 was grown without chlorophenol for two successive transfers. The last culture was used to inoculate three cultures (500 ml) without chlorophenol. After an incubation at 30°C for 30 h (optical density at 600 nm [OD 600 ], 0.25 to 0.27), the first culture received 2,4,6-TCP at 50 M (final concentration) and the second 3,5-DCP at 60 M (final concentration). No chlorophenol was added to the third culture (unexposed culture). Aliquots of the cultures (20 ml) were transferred into 70-ml sterile serum bottles. After various incubation periods (0,4,8,18, 28, and 44 h) at 30°C, a 2-ml sample was collected for measurin...

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Divergent roles of CprK paralogues from Desulfitobacterium hafniense in activating gene expression

Cited by 20 publications

References 42 publications

Distribution of dehalogenation activity in subseafloor sediments of the Nankai Trough subduction zone

Distribution of dehalogenation activity in subseafloor sediments of the Nankai Trough subduction zone

The restricted metabolism of the obligate organohalide respiring bacterium Dehalobacter restrictus: lessons from tiered functional genomics

Quantitative Analysis of the Relative Transcript Levels of Four Chlorophenol Reductive Dehalogenase Genes in Desulfitobacterium hafniense PCP-1 Exposed to Chlorophenols

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