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Signature Arsenic Detoxification Pathways in Halomonas sp. Strain GFAJ-1.

ABSTRACT: Since the original report that Halomonas sp. strain GFAJ-1 was capable of using arsenic instead of phosphorus to sustain growth, additional studies have been conducted, and GFAJ-1 is now considered a highly arsenic-resistant but phosphorus-dependent bacterium. However, the mechanisms supporting the extreme arsenic resistance of the GFAJ-1 strain remain unknown. In this study, we show that GFAJ-1 has multiple distinct arsenic resistance mechanisms. It lacks the genes to reduce arsenate, which is the essential step in the well-characterized resistance mechanism of arsenate reduction coupled to arsenite extrusion. Instead, GFAJ-1 has two arsenic resistance operons, arsH1-acr3-2-arsH2 and mfs1-mfs2-gapdh, enabling tolerance to high levels of arsenate. mfs2 and gapdh encode proteins homologous to Pseudomonas aeruginosa ArsJ and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), respectively, which constitute the equivalent of an As(V) efflux system to catalyze the transformation of inorganic arsenate to pentavalent organoarsenical 1-arseno-3-phosphoglycerate and its subsequent extrusion. Surprisingly, the arsH1-acr3-2-arsH2 operon seems to consist of typical arsenite resistance genes, but this operon is sufficient to confer both arsenite and arsenate resistance on Escherichia coli AW3110 even in the absence of arsenate reductase, suggesting a novel pathway of arsenic detoxification. The simultaneous occurrence of these two unusual detoxification mechanisms enables the adaptation of strain GFAJ-1 to the particularly arsenic-rich environment of Mono Lake.IMPORTANCEHalomonas sp. strain GFAJ-1 was previously reported to use arsenic as a substitute for phosphorus to sustain life under phosphate-limited conditions. Although this claim was later undermined by several groups, how GFAJ-1 can thrive in environments with high arsenic concentrations remains unclear. Here, we determined that this ability can be attributed to the possession of two arsenic detoxification operons, arsH1-acr3-2-arsH2 and mfs1-mfs2-gapdhmfs2 and gapdh encode proteins homologous to ArsJ and GAPDH in Pseudomonas aeruginosa; these proteins create an arsenate efflux pathway to reduce cellular arsenate accumulation. Interestingly, the combination of acr3-2 with either arsH gene was sufficient to confer resistance to both arsenite and arsenate in E. coli AW3110, even in the absence of arsenate reductase, suggesting a new strategy for bacterial arsenic detoxification. This study concludes that the survival of GFAJ-1 in high arsenic concentrations is attributable to the cooccurrence of these two unusual arsenic detoxification mechanisms.

SUBMITTER: Wu S

PROVIDER: S-EPMC5930303 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Signature Arsenic Detoxification Pathways in <i>Halomonas</i> sp. Strain GFAJ-1.

Wu Shuangju S Wang Lianrong L Gan Rui R Tong Tong T Bian Hao H Li Zhiqiang Z Du Shiming S Deng Zixin Z Chen Shi S

mBio 20180501 3

Since the original report that <i>Halomonas</i> sp. strain GFAJ-1 was capable of using arsenic instead of phosphorus to sustain growth, additional studies have been conducted, and GFAJ-1 is now considered a highly arsenic-resistant but phosphorus-dependent bacterium. However, the mechanisms supporting the extreme arsenic resistance of the GFAJ-1 strain remain unknown. In this study, we show that GFAJ-1 has multiple distinct arsenic resistance mechanisms. It lacks the genes to reduce arsenate, wh ...[more]

PMID: 29717010

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Project description:Insight into the mechanisms for arsenic detoxification by Geobacter species is expected to improve the understanding of global cycling of arsenic in iron-rich subsurface sedimentary environments. Analysis of 14 different Geobacter genomes showed that all of these species have genes coding for an arsenic detoxification system (ars operon), and several have genes required for arsenic respiration (arr operon) and methylation (arsM). Genes encoding four arsenic repressor-like proteins were detected in the genome of G. sulfurreducens; however, only one (ArsR1) regulated transcription of the ars operon. Elimination of arsR1 from the G. sulfurreducens chromosome resulted in enhanced transcription of genes coding for the arsenic efflux pump (Acr3) and arsenate reductase (ArsC). When the gene coding for Acr3 was deleted, cells were not able to grow in the presence of either the oxidized or reduced form of arsenic, while arsC deletion mutants could grow in the presence of arsenite but not arsenate. These studies shed light on how Geobacter influences arsenic mobility in anoxic sediments and may help us develop methods to remediate arsenic contamination in the subsurface. This study examines arsenic transformation mechanisms utilized by Geobacter, a genus of iron-reducing bacteria that are predominant in many anoxic iron-rich subsurface environments. Geobacter species play a major role in microbially mediated arsenic release from metal hydroxides in the subsurface. This release raises arsenic concentrations in drinking water to levels that are high enough to cause major health problems. Therefore, information obtained from studies of Geobacter should shed light on arsenic cycling in iron-rich subsurface sedimentary environments, which may help reduce arsenic-associated illnesses. These studies should also help in the development of biosensors that can be used to detect arsenic contaminants in anoxic subsurface environments. We examined 14 different Geobacter genomes and found that all of these species possess genes coding for an arsenic detoxification system (ars operon), and some also have genes required for arsenic respiration (arr operon) and arsenic methylation (arsM).

Project description:Two NhaD-type antiporters, NhaD1 and NhaD2, from the halotolerant and alkaliphilic Halomonas sp. strain Y2, exhibit different physiological functions in regard to Na+ and Li+ resistance, although they share high sequence identity. In the present study, the truncation of an additional 4 C-terminal residues from NhaD2 or an exchange of 39 N-terminal residues between these proteins resulted in the complete loss of antiporter activity. Interestingly, combining 39 N-terminal residues and 7 C-terminal residues of NhaD2 (N39D2-C7) partially recovered the activity for Na+ and Li+ expulsion, as well as complementary growth following exposure to 300 mM Na+ and 100 mM Li+ stress. The recovered activity of chimera N39D2-C7 indicated that the N and C termini are structurally dependent on each other and function synergistically. Furthermore, fluorescence resonance energy transfer (FRET) analysis suggested that the N and C termini are relatively close in proximity which may account for their synergistic function in ion translocation. In the N-terminal region of N39D2-C7, the replacement of Glu38 with Pro abolished the recovered complementary and transport activities. In addition, this amino acid substitution in NhaD2 resulted in a drastically decreased complementation ability in Escherichia coli KNabc (level identical to that of NhaD1), as well as decreased activity and an altered pH profile.IMPORTANCE Limited information on NhaD antiporters supports speculation that these antiporters are important for resistance to high salinity and alkalinity. Moreover, only a few functional residues have been identified in NhaD antiporters, and there is limited literature on the molecular mechanisms of NhaD antiporter activity. The altered antiporter abilities of chimeras and mutants in this study implicate the functions of the N and C termini, especially Glu38, in pH regulation and ion translocation, and, most importantly, the essential roles of this negatively charged residue in maintaining the physiological function of NhaD2. These findings further our understanding of the molecular mechanism of NhaD antiporters for ion transport.

Dataset Information

Signature Arsenic Detoxification Pathways in Halomonas sp. Strain GFAJ-1.

Publications

Signature Arsenic Detoxification Pathways in <i>Halomonas</i> sp. Strain GFAJ-1.

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