Efflux Transporter ArsK Is Responsible for Bacterial Resistance to Arsenite, Antimonite, Trivalent Roxarsone, and Methylarsenite

Abstract

Arsenic-resistant bacteria have evolved various efflux systems for arsenic resistance. Five arsenic efflux proteins, ArsB, Acr3, ArsP, ArsJ, and MSF1, have been reported. In this study, comprehensive analyses were performed to study the function of a putative major facilitator superfamily gene, arsK, and the regulation of arsK transcriptional expression in Agrobacterium tumefaciens GW4. We found that (i) arsK is located on an arsenic gene island in strain GW4. ArsK orthologs are widely distributed in arsenic-resistant bacteria and are phylogenetically divergent from the five reported arsenic efflux proteins, indicating that it may be a novel arsenic efflux transporter. (ii) Reporter gene assays showed that the expression of arsK was induced by arsenite [As(III)], antimonite [Sb(III)], trivalent roxarsone [Rox(III)], methylarsenite [MAs(III)], and arsenate [As(V)]. (iii) Heterologous expression of ArsK in an arsenic-hypersensitive Escherichia coli strain showed that ArsK was essential for resistance to As(III), Sb(III), Rox(III), and MAs(III) but not to As(V), dimethylarsenite [dimethyl-As(III)], or Cd(II). (iv) ArsK reduced the cellular accumulation of As(III), Sb(III), Rox(III), and MAs(III) but not to As(V) or dimethyl-As(III). (v) A putative arsenic regulator gene arsR2 was cotranscribed with arsK, and (vi) ArsR2 interacted with the arsR2-arsK promoter region without metalloids and was derepressed by As(III), Sb(III), Rox(III), and MAs(III), indicating the repression activity of ArsR2 for the transcription of arsK. These results demonstrate that ArsK is a novel arsenic efflux protein for As(III), Sb(III), Rox(III), and MAs(III) and is regulated by ArsR2. Bacteria use the arsR2-arsK operon for resistance to several trivalent arsenicals or antimonials. IMPORTANCE The metalloid extrusion systems are very important bacterial resistance mechanisms. Each of the previously reported ArsB, Acr3, ArsP, ArsJ, and MSF1 transport proteins conferred only inorganic or organic arsenic/antimony resistance. In contrast, ArsK confers resistance to several inorganic and organic trivalent arsenicals and antimonials. The identification of the novel efflux transporter ArsK enriches our understanding of bacterial resistance to trivalent arsenite [As(III)], antimonite [Sb(III)], trivalent roxarsone [Rox(III)], and methylarsenite [MAs(III)].

The antibiotic action of methylarsenite is an emergent property of microbial communities

“…Both ArsM and ArsP are found in anaerobes, consistent with an early co‐evolution in an anoxic environment (Chen et al , ). Quite recently, an additional efflux permease for As(III), Sb(III), Rox(III) and MAs(III) that confers resistance to those arsenicals was identified in an ars operon from Agrobacterium tumefaciens GW4 (Shi et al , ). ArsK is widely distributed in bacteria but not in archaea, fungi, plants or animals.…”

Arsenic and the gastrointestinal tract microbiome

“…Acr3 is another extrusion mechanism and its encoding gene is actually found more frequently than arsB in ars operons (Yang et al, 2015). Just recently, Shi et al (2018) characterized ArsK, which also facilitates extrusion of As(III) as well as other trivalent metalloids (Fig. 3).…”

“…Just recently, Shi et al . () characterized ArsK, which also facilitates extrusion of As(III) as well as other trivalent metalloids (Fig. ).…”

The Arsenic Methylation Cycle: How Microbial Communities Adapted Methylarsenicals for Use as Weapons in the Continuing War for Dominance

“…Recently a second MAs(III) efflux permease, ArsK, was found in an arsenic gene island in the chromosome of Agrobacterium tumefaciens GW4 (Shi et al, 2018) as well as in a Bacillus sp. strain (Jia et al, 2019).…”

“…Life has not only adapted effectively to their own environments enrich with arsenic, but also can utilize arsenic as an offensive weapon to gain a competitive advantage over sensitive members of the complex microbial communities (Chen et al, 2019b). The original antibiotic producer (ArsM hosts) may have not only defended itself by extrusion of its own toxic products MAs(III) with methylarsenite efflux permeases (ArsP or ArsK) (Chen et al, 2015b;Shi et al, 2018) but also put the toxin outside of the cells, where it could kill its sensitive neighbors (Figure 1). There is some evidence that suggests that arsP usually found to co-exists with arsM, primarily in anaerobes, which might be an ancestral feature in lineages of anaerobes.…”