Factors influencing the diversity of iron uptake systems in aquatic microorganisms

Abstract

Iron (Fe) is an essential micronutrient for many processes in all living cells. Dissolved Fe (dFe) concentrations in the ocean are of the order of a few nM, and Fe is often a factor limiting primary production. Bioavailability of Fe in aquatic environments is believed to be primarily controlled through chelation by Fe-binding ligands. Marine microbes have evolved different mechanisms to cope with the scarcity of bioavailable dFe. Gradients in dFe concentrations and diversity of the Fe-ligand pool from coastal to open ocean waters have presumably imposed selection pressures that should be reflected in the genomes of microbial communities inhabiting the pelagic realm. We applied a hidden Markov model (HMM)-based search for proteins related to cellular iron metabolism, and in particular those involved in Fe uptake mechanisms in 164 microbial genomes belonging to diverse taxa and occupying different aquatic niches. A multivariate statistical approach demonstrated that in phototrophic organisms, there is a clear influence of the ecological niche on the diversity of Fe uptake systems. Extending the analyses to the metagenome database from the Global Ocean Sampling expedition, we demonstrated that the Fe uptake and homeostasis mechanisms differed significantly across marine niches defined by temperatures and dFe concentrations, and that this difference was linked to the distribution of microbial taxa in these niches. Using the dN/dS ratios (which signify the rate of non-synonymous mutations) of the nucleotide sequences, we identified that genes encoding for TonB, Ferritin, Ferric reductase, IdiA, ZupT, and Fe(2+) transport proteins FeoA and FeoB were evolving at a faster rate (positive selection pressure) while genes encoding ferrisiderophore, heme and Vitamin B12 uptake systems, siderophore biosynthesis, and IsiA and IsiB were under purifying selection pressure (evolving slowly).

Keywords: Fe limitation; Fe- binding ligands; aquatic niches; dN/dS ratio; eukaryotic phytoplankton; marine microbes; metagenomes; multivariate statistics.

Figure 1

Distribution of components of Fe-metabolism systems in (A) marine microbial taxa and (B) Temperature groupings of metagenomes. For detailed description of the components of each system see Table 1.

Figure 2

Phylogenetic trees of (A) RhbA, (B) RhbB, and (C) RhtX protein sequences from Open Ocean picocyanobacteria and eukaryotic phytoplankton recovered in our study. The tree was constructed using a Neighbor-Joining method, using the Jukes–Cantor correction and a bootstrap test was conducted with 1000 replicates. The scale bar represents 100% estimated sequence divergence.

Figure 3

Non-parametric multidimensional scaling plots showing the clustering of genomes and metagenomes based on the similarities of occurrence patterns of Fe uptake systems. (A) Heterotrophic genomes labeled by taxa, (B) phototrophic genomes labeled by taxa, (C) phototrophic genomes labeled by niche, and (D) GOS metagenomes labeled by marine niche groups.

Figure 4

Coordinates of GOS metagenome samples overlayed on dissolved Fe concentrations as predicted by PELAGOS model.

Figure 5

Principal Components Analysis of GOS samples using similarities of the environmental parameter profiles. The samples are labeled according to the ocean basins (Atlantic or Pacific) combined with niche groups (Open Ocean or Coastal).

Figure 6

Median and range of dissolved Fe concentrations (A) and Temperature (B) for the GOS metagenome sample groups used in the study. Numbers of samples in the groups are as follows: Atlantic (18), Pacific (12), Atlantic Open Ocean (9), Pacific Open Ocean (5), Pacific-Coastal (7), Tropical-Open Ocean (11), Tropical-Coastal (11), Temperate-Coastal (8), Open Ocean (14), Coastal (16).

Figure 7

(A) Average dN/dS ratios of selected genes. The genes are sorted by highest dN/dS value. (B) Phylogenetic spread as defined by maximum phylogenetic distance among the genomes possessing the gene. The black line marks the dN/dS value = 1. Genes with dN/dS >1 and a wide phylogenetic spread are marked with stars. Genes with dN/dS >1 and a narrow phylogenetic spread are marked by an arrow. Fe-red – Ferric reductase.