Actinorhodopsins: proteorhodopsin‐like gene sequences found predominantly in non‐marine environments

Abstract

Proteorhodopsins are light-energy-harvesting transmembrane proteins encoded by genes recently discovered in the surface waters of the world's oceans. Metagenomic data from the Global Ocean Sampling expedition (GOS) recovered 2674 proteorhodopsin-related sequences from 51 aquatic samples. Four of these samples were from non-marine environments, specifically, Lake Gatun within the Panama Canal, Delaware Bay and Chesapeake Bay and the Punta Cormorant Lagoon in Ecuador. Rhodopsins related to but phylogenetically distinct from most sequences designated proteorhodopsins were present at all four of these non-marine sites and comprised three different clades that were almost completely absent from marine samples. Phylogenomic analyses of genes adjacent to those encoding these novel rhodopsins suggest affiliation to the Actinobacteria, and hence we propose to name these divergent, non-marine rhodopsins 'actinorhodopsins'. Actinorhodopsins conserve the acidic amino acid residues critical for proton pumping and their genes lack genomic association with those encoding photo-sensory transducer proteins, thus supporting a putative ion pumping function. The ratio of recA and radA to rhodopsin genes in the different environment types sampled within the GOS indicates that rhodopsins of one type or another are abundant in microbial communities in freshwater, estuarine and lagoon ecosystems, supporting an important role for these photosystems in all aquatic environments influenced by sunlight.

Actinorhodopsin genes discovered in diverse freshwater habitats and among cultivated freshwater Actinobacteria

“…Our earlier analyses also consisted of a bioinformatic assessment of GOS-derived (Rusch et al, 2007) metagenomic assemblies containing rhodopsin genes of these three novel clades and found they were almost exclusively linked to open reading frames most closely related to homologues encoded in sequenced actinobacterial genomes. In Sharma et al (2008) we designated these novel clades of rhodopsins collectively as 'actinorhodopsins (ActRs)' based on (1) the aforementioned pattern of gene linkage on GOS metagenomic assemblies, (2) the observation by Rusch et al (2007) that actinobacterial SSU rRNA sequences were abundant in non-marine samples and (3) the knowledge that Actinobacteria are common inhabitants of freshwater ecosystems (Warnecke et al, 2004). However in our previous study, we were unable to identify the specific actinobacterial carriers of ActRs from GOS samples.…”

“…Biomass was collected from the central basin of Lake Erie (Environment Canada Station 84, 411 56 0 46 00 N, 811 38 0 46 00 W) during a July 2005 cruise aboard the CCGS Limnos when the water column was thermally stratified (for sampling details see Wilhelm et al, 2006). DNA for rhodopsin PCR was integrated from Sharma et al (2008), we described three clades of actinorhodopsins (ActR) identified from the Global Ocean Sampling (GOS) expedition's metagenomic samples of non-marine sites (Rusch et al, 2007). In addition to ActR sequences from our earlier study, this tree also contains five full-length LG1 ActR sequences obtained here, from actinobacterial cultures MWH-Uga1, MWH-Dar1, MWH-Dar4, MWH-Ta8, MWH-EgelM2-3.…”

“…However in our previous study, we were unable to identify the specific actinobacterial carriers of ActRs from GOS samples. On the basis of our findings in Sharma et al (2008), we predicted that ActR gene sequences would likely be found in other freshwater environments and in the genomes of cultivated Actinobacteria. To test these predictions, we designed degenerate primers and used the polymerase chain reaction (PCR) to amplify ActR genes from freshwater metagenomic DNA samples and Actinobacteria cultivated from various freshwater sources.…”

“…Degenerate PCR primers (Table 1) were designed to amplify partial gene sequences from ActR sequences clades (LG1, LG2 and PCL1) uncovered from our previous study (Sharma et al, 2008). Rhodopsins were amplified from metagenomic DNA samples using either the combined set of three primers targeting ActR clades LG1 and LG2 (Table 1) or a set of three degenerate primers previously designed by Sabehi et al (2005) to target proteorhodopsin.…”

“…Rusch et al described the presence of these genes among microbes inhabiting the freshwater Lake Gatun (part of the Panama Canal), the estuaries Delaware Bay and Chesapeake Bay, and the hypersaline Punta Cormorant lagoon in Ecuador. In a separate analysis of rhodopsin sequences from these non-marine environments in Sharma et al (2008), we described three novel groups of sequences (LG1, LG2 and PCL1, Figure 1) comprising the majority of rhodopsin diversity. Our earlier analyses also consisted of a bioinformatic assessment of GOS-derived (Rusch et al, 2007) metagenomic assemblies containing rhodopsin genes of these three novel clades and found they were almost exclusively linked to open reading frames most closely related to homologues encoded in sequenced actinobacterial genomes.…”