New findings on evolution of metal homeostasis genes: evidence from comparative genome analysis of bacteria and archaea

Abstract

In order to examine the natural history of metal homeostasis genes in prokaryotes, open reading frames with homology to characterized P(IB)-type ATPases from the genomes of 188 bacteria and 22 archaea were investigated. Major findings were as follows. First, a high diversity in N-terminal metal binding motifs was observed. These motifs were distributed throughout bacterial and archaeal lineages, suggesting multiple loss and acquisition events. Second, the CopA locus separated into two distinct phylogenetic clusters, CopA1, which contained ATPases with documented Cu(I) influx activity, and CopA2, which contained both efflux and influx transporters and spanned the entire diversity of the bacterial domain, suggesting that CopA2 is the ancestral locus. Finally, phylogentic incongruences between 16S rRNA and P(IB)-type ATPase gene trees identified at least 14 instances of lateral gene transfer (LGT) that had occurred among diverse microbes. Results from bootstrapped supported nodes indicated that (i) a majority of the transfers occurred among proteobacteria, most likely due to the phylogenetic relatedness of these organisms, and (ii) gram-positive bacteria with low moles percent G+C were often involved in instances of LGT. These results, together with our earlier work on the occurrence of LGT in subsurface bacteria (J. M. Coombs and T. Barkay, Appl. Environ. Microbiol. 70:1698-1707, 2004), indicate that LGT has had a minor role in the evolution of P(IB)-type ATPases, unlike other genes that specify survival in metal-stressed environments. This study demonstrates how examination of a specific locus across microbial genomes can contribute to the understanding of phenotypes that are critical to the interactions of microbes with their environment.

FIG. 1.

Amino acid sequence alignments showing the different conserved regions of the N termini of P_IB-type ATPases. Amino acids specific to the motif are boxed in black, while amino acids that align but are not specific to the motif are boxed in gray. (A) Alignment of sequences from selected Gammaproteobacteria demonstrating the consensus metal binding motif (GMXCXXC). (B) Alignment of sequences from selected gram-positive bacteria demonstrating the histidine-rich metal binding motif. (C) Alignment of sequences from selected gram-positive bacteria showing lack of any known metal binding motif.

FIG. 2.

Phylogeny of ZntA-like sequences from bacterial genomes. Neighbor-joining analysis of sequences from completed bacterial genomes that were identified by their homology to ZntA-like P_IB-type ATPases is shown. Circles at each node indicate the level of bootstrap support obtained with both parsimony and distance methods: black, >80%; gray, >50%; white, supported at >50% by one method only. Dark and light gray boxes indicate the presence of a histidine-rich metal binding motif or no metal binding motif at the N terminus, respectively. White boxes indicate both CXXC and histidine-rich motifs. Black ovals mark sequences that demonstrate phylogenetic incongruence compared to the 16S rRNA phylogeny.

FIG. 3.

Phylogeny of CopA-like sequences from bacterial genomes. These sequences were identified on the basis of sequence similarity to CopA-like P_IB-type ATPases. See the Fig. 2 legend for details.

FIG. 4.

Phylogeny of ZntA-like and CopA-like sequences from archaeal genomes. (A) Neighbor-joining analysis of 16S rRNA gene (left) and P_IB-type ATPase (right) sequences. Circles at each node indicate the level of bootstrap support obtained when analysis was by both parsimony and distance methods (see the legend to Fig. 2). Instances of incongruence between the two trees are indicated by black ovals. (B) Neighbor-joining analysis of select ZntA-like P_IB-type ATPases belonging to both the bacterial and archaeal domains. Evidence supporting a single interdomain gene transfer is boxed and marked with a black oval.