title: Role of an archaeal PitA transporter in the copper and arsenic resistance of Metallosphaera sedula, an extreme thermoacidophile.
abstract: Thermoacidophilic archaea, such as Metallosphaera sedula, are lithoautotrophs that occupy metal-rich environments. In previous studies, an M. sedula mutant lacking the primary copper efflux transporter, CopA, became copper sensitive. In contrast, the basis for supranormal copper resistance remained unclear in the spontaneous M. sedula mutant, CuR1. Here, transcriptomic analysis of copper-shocked cultures indicated that CuR1 had a unique regulatory response to metal challenge corresponding to the upregulation of 55 genes. Genome resequencing identified 17 confirmed mutations unique to CuR1 that were likely to change gene function. Of these, 12 mapped to genes with annotated function associated with transcription, metabolism, or transport. These mutations included 7 nonsynonymous substitutions, 4 insertions, and 1 deletion. One of the insertion mutations mapped to pseudogene Msed_1517 and extended its reading frame an additional 209 amino acids. The extended mutant allele was identified as a homolog of Pho4, a family of phosphate symporters that includes the bacterial PitA proteins. Orthologs of this allele were apparent in related extremely thermoacidophilic species, suggesting M. sedula naturally lacked this gene. Phosphate transport studies combined with physiologic analysis demonstrated M. sedula PitA was a low-affinity, high-velocity secondary transporter implicated in copper resistance and arsenate sensitivity. Genetic analysis demonstrated that spontaneous arsenate-resistant mutants derived from CuR1 all underwent mutation in pitA and nonselectively became copper sensitive. Taken together, these results point to archaeal PitA as a key requirement for the increased metal resistance of strain CuR1 and its accelerated capacity for copper bioleaching.
authors.affiliation: Beadle Center for Genetics, University of Nebraska, Lincoln, Nebraska, USA.
authors.forename: Samuel
authors.initials: S
authors.lastname: McCarthy
authors.affiliation: Beadle Center for Genetics, University of Nebraska, Lincoln, Nebraska, USA School of Minerals Processing and Bioengineering, Central South University, Changsha, China.
authors.forename: Chenbing
authors.initials: C
authors.lastname: Ai
authors.affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, North Carolina, USA.
authors.forename: Garrett
authors.initials: G
authors.lastname: Wheaton
authors.affiliation: Beadle Center for Genetics, University of Nebraska, Lincoln, Nebraska, USA.
authors.forename: Rahul
authors.initials: R
authors.lastname: Tevatia
authors.affiliation: Beadle Center for Genetics, University of Nebraska, Lincoln, Nebraska, USA.
authors.forename: Valerie
authors.initials: V
authors.lastname: Eckrich
authors.affiliation: Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, North Carolina, USA.
authors.forename: Robert
authors.initials: R
authors.lastname: Kelly
authors.affiliation: Beadle Center for Genetics, University of Nebraska, Lincoln, Nebraska, USA pblum1@unl.edu.
authors.forename: Paul
authors.initials: P
authors.lastname: Blum
mesh_terms: D019843:Archaeal Proteins / Q000235:genetics / Q000378:metabolism*; D001151:Arsenic / Q000378:metabolism / Q000633:toxicity*; D001692:Biological Transport; D003300:Copper / Q000378:metabolism / Q000633:toxicity*; D019848:Gene Expression Regulation, Archaeal / Q000502:physiology*; D020745:Genome, Archaeal; D008969:Molecular Sequence Data; D016943:Sulfolobaceae / Q000187:drug effects* / Q000378:metabolism*
chemical_list: D019843:Archaeal Proteins; D003300:Copper; D001151:Arsenic
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