Gazalah Sabehi

Diversification and spectral tuning in marine proteorhodopsins

Proteorhodopsins, ubiquitous retinylidene photoactive proton pumps, were recently discovered in the cosmopolitan uncultured SAR86 bacterial group in oceanic surface waters. Two related proteorhodopsin families were found that absorb light with different absorption maxima, 525 nm (green) and 490 nm (blue), and their distribution was shown to be stratified with depth. Using structural modeling comparisons and mutagenesis, we report here on a single amino acid residue at position 105 that functions as a spectral tuning switch and accounts for most of the spectral difference between the two pigment families. Furthermore, looking at natural environments, we found novel proteorhodopsin gene clusters spanning the range of 540–505 nm and containing changes in the same identified key switch residue leading to changes in their absorption maxima. The results suggest a simultaneous diversification of green proteorhodopsin and the new key switch variant pigments. Our observations demonstrate that this single‐residue switch mechanism is the major determinant of proteorhodopsin wavelength regulation in natural marine environments.

New Insights into Metabolic Properties of Marine Bacteria Encoding Proteorhodopsins

Proteorhodopsin phototrophy was recently discovered in oceanic surface waters. In an effort to characterize uncultured proteorhodopsin-exploiting bacteria, large-insert bacterial artificial chromosome (BAC) libraries from the Mediterranean Sea and Red Sea were analyzed. Fifty-five BACs carried diverse proteorhodopsin genes, and we confirmed the function of five. We calculate that proteorhodopsin-exploiting bacteria account for 13% of microorganisms in the photic zone. We further show that some proteorhodopsin-containing bacteria possess a retinal biosynthetic pathway and a reverse sulfite reductase operon, employed by prokaryotes oxidizing sulfur compounds. Thus, these novel phototrophs are an unexpectedly large and metabolically diverse component of the marine microbial surface water.

Widespread distribution of proteorhodopsins in freshwater and brackish ecosystems

Proteorhodopsins (PRs) are light-driven proton pumps that have been found in a variety of marine environments. The goal of this study was to search for PR presence in different freshwater and brackish environments and to explore the diversity of non-marine PR protein. Here, we show that PRs exist in distinctly different aquatic environments, ranging from clear water lakes to peat lakes and in the Baltic Sea. Some of the PRs observed in this study formed unique clades that were not previously observed in marine environments, whereas others were similar to PRs found in non-marine samples of the Global Ocean Sampling (GOS) expedition. Furthermore, the similarity of several PRs isolated from lakes in different parts of the world suggests that these genes are dispersed globally and that they may encode unique functional capabilities enabling successful competition in a wide range of freshwater environments. Phylogenomic analysis of genes found on these GOS scaffolds suggests that some of the freshwater PRs are found in freshwater Flavobacteria and freshwater SAR11-like bacteria.

Roseobacter-Like Bacteria in Red and Mediterranean Sea Aerobic Anoxygenic Photosynthetic Populations

ABSTRACT Bacteriochlorophyll a-containing aerobic anoxygenic phototrophs (AAnP) have been proposed to account for up to 11% of the total surface water microbial community and to potentially have great ecological importance in the worlds oceans. Recently, environmental and genomic data based on analysis of the pufM gene identified the existence of α-proteobacteria as well as possible γ-like proteobacteria among AAnP in the Pacific Ocean. Here we report on analyses of environmental samples from the Red and Mediterranean Seas by using pufM as well as the bchX and bchL genes as molecular markers. The majority of photosynthesis genes retrieved from these seas were related to Roseobacter-like AAnP sequences. Furthermore, the sequence of a novel photosynthetic operon organization from an uncultured Roseobacter-like bacterial artificial chromosome retrieved from the Red Sea is described. The data show the presence of Roseobacter-like bacteria in Red and Mediterranean Sea AAnP populations in the seasons analyzed.

Adaptation and spectral tuning in divergent marine proteorhodopsins from the eastern Mediterranean and the Sargasso Seas

Proteorhodopsins (PRs) phototrophy was recently discovered in oceanic surface waters. PRs have been observed in different marine environments and in diverse taxa, including the ubiquitous marine alphaproteobacterial SAR11 group and the uncultured gammaproteobacterial SAR86 group. Previously, two SAR86 PR subgroups, discovered in the Pacific Ocean, were shown to absorb light with different maxima, λmax 527 nm (green) and λmax 490 nm (blue) and their distribution was explained by prevailing light conditions – green pigments at the surface and blue in deeper waters. Here, we show that PRs display high diversity in geographically distinct patterns despite similar physical water column properties such as mixing and light penetration. We compared summer and winter samples representing stratified and mixed conditions from both the Mediterranean and Sargasso Sea. As expected, in the Mediterranean Sea, green pigments were mainly confined to the surface and the percentage of blue pigments increased toward deeper samples; in the Sargasso Sea, unexpectedly, all PRs were of the blue type. As an additional result, both locations show seasonal dependence in the distribution of different PR families. Finally, spectral tuning was not restricted to a single PR family as previously reported but occurs across the sampled PR families from various microbial taxa. The distribution of tunable PRs across the PR tree suggests that ready adaptability has been distributed widely among microorganisms, and may be a reason that PRs are abundant and taxonomically widely dispersed.

A novel lineage of myoviruses infecting cyanobacteria is widespread in the oceans

Viruses infecting bacteria (phages) are thought to greatly impact microbial population dynamics as well as the genome diversity and evolution of their hosts. Here we report on the discovery of a novel lineage of tailed dsDNA phages belonging to the family Myoviridae and describe its first representative, S-TIM5, that infects the ubiquitous marine cyanobacterium, Synechococcus. The genome of this phage encodes an entirely unique set of structural proteins not found in any currently known phage, indicating that it uses lineage-specific genes for virion morphogenesis and represents a previously unknown lineage of myoviruses. Furthermore, among its distinctive collection of replication and DNA metabolism genes, it carries a mitochondrial-like DNA polymerase gene, providing strong evidence for the bacteriophage origin of the mitochondrial DNA polymerase. S-TIM5 also encodes an array of bacterial-like metabolism genes commonly found in phages infecting cyanobacteria including photosynthesis, carbon metabolism and phosphorus acquisition genes. This suggests a common gene pool and gene swapping of cyanophage-specific genes among different phage lineages despite distinct sets of structural and replication genes. All cytosines following purine nucleotides are methylated in the S-TIM5 genome, constituting a unique methylation pattern that likely protects the genome from nuclease degradation. This phage is abundant in the Red Sea and S-TIM5 gene homologs are widespread in the oceans. This unusual phage type is thus likely to be an important player in the oceans, impacting the population dynamics and evolution of their primary producing cyanobacterial hosts.

Microbial community genomics in eastern Mediterranean Sea surface waters

Offshore waters of the eastern Mediterranean Sea are one of the most oligotrophic regions on Earth in which the primary productivity is phosphorus limited. To study the unexplored function and physiology of microbes inhabiting this system, we have analyzed a genomic library from the eastern Mediterranean Sea surface waters by sequencing both termini of nearly 5000 clones. Genome recruitment strategies showed that the majority of high-scoring pairs corresponded to genomes from the Alphaproteobacteria (SAR11-like and Rhodobacterales), Cyanobacteria (Synechococcus and high-light adapted Prochlorococcus) and diverse uncultured Gammaproteobacteria. The community structure observed, as evaluated by both protein similarity scores or metabolic potential, was similar to that found in the euphotic zone of the ALOHA station off Hawaii but very different from that of deep aphotic zones in both the Mediterranean Sea and the Pacific Ocean. In addition, a strong enrichment toward phosphate and phosphonate uptake and utilization metabolism was also observed.

Diversity and evolutionary relationships of T7-like podoviruses infecting marine cyanobacteria

Phages are extremely abundant in the oceans, influencing the population dynamics, diversity and evolution of their hosts. Here we assessed the diversity and phylogenetic relationships among T7-like cyanophages using DNA polymerase (replication), major capsid (structural) and photosynthesis psbA (host-derived) genes from isolated phages. DNA polymerase and major capsid phylogeny divided them into two discrete clades with no evidence for gene exchange between clades. Clade A phages primarily infect Synechococcus while clade B phages infect either Synechococcus or Prochlorococcus. The major capsid gene of one of the phages from clade B carries a putative intron. Nearly all clade B phages encode psbA whereas clade A phages do not. This suggests an ancient separation between cyanophages from these two clades, with the acquisition or loss of psbA occurring around the time of their divergence. A mix and match of clustering patterns was found for the replication and structural genes within each major clade, even among phages infecting different host genera. This is suggestive of numerous gene exchanges within each major clade and indicates that core phage functions have not coevolved with specific hosts. In contrast, clustering of phage psbA broadly tracks that of the host genus. These findings suggest that T7-like cyanophages evolve through clade-limited gene exchanges and that different genes are subjected to vastly different selection pressures.

Transcriptome dynamics of a broad host-range cyanophage and its hosts

Cyanobacteria are highly abundant in the oceans and are constantly exposed to lytic viruses. The T4-like cyanomyoviruses are abundant in the marine environment and have broad host-ranges relative to other cyanophages. It is currently unknown whether broad host-range phages specifically tailor their infection program for each host, or employ the same program irrespective of the host infected. Also unknown is how different hosts respond to infection by the same phage. Here we used microarray and RNA-seq analyses to investigate the interaction between the Syn9 T4-like cyanophage and three phylogenetically, ecologically and genomically distinct marine Synechococcus strains: WH7803, WH8102 and WH8109. Strikingly, Syn9 led a nearly identical infection and transcriptional program in all three hosts. Different to previous assumptions for T4-like cyanophages, three temporally regulated gene expression classes were observed. Furthermore, a novel regulatory element controlled early-gene transcription, and host-like promoters drove middle gene transcription, different to the regulatory paradigm for T4. Similar results were found for the P-TIM40 phage during infection of Prochlorococcus NATL2A. Moreover, genomic and metagenomic analyses indicate that these regulatory elements are abundant and conserved among T4-like cyanophages. In contrast to the near-identical transcriptional program employed by Syn9, host responses to infection involved host-specific genes primarily located in hypervariable genomic islands, substantiating islands as a major axis of phage–cyanobacteria interactions. Our findings suggest that the ability of broad host-range phages to infect multiple hosts is more likely dependent on the effectiveness of host defense strategies than on differential tailoring of the infection process by the phage.

Comparative analyses of actinobacterial genomic fragments from Lake Kinneret

The high genomic G+C group of Actinobacteria possesses a variety of physiological and metabolic properties, and exhibits diverse lifestyles and ecological distribution. In recent years, Actinobacteria have been found to frequently dominate samples obtained from freshwater samples. Furthermore, phylogenetic analyses have shown that 16S rRNA genes from uncultured actinobacterial freshwater samples cluster in four distinct lineages. While these lineages are abundant, little is known about them and currently no pure-culture representatives or genomic fragments of them are available. In a screen of a genomic library from the moderately eutrophic freshwater Lake Kinneret, five fosmid clones containing actinobacterial genomic fragments were found. Three approximately 40 kb genomic fragments were chosen for sequencing. Fosmids K003 and K005 showed high similarity and were affiliated with the acIV actinobacterial freshwater lineage. Fosmid K004 was affiliated with the highly abundant acI lineage. A comparative genomic analysis revealed high synteny between the two freshwater clones K003 and K005 but a lower synteny between these two and the K004 fosmid. Fosmids K003 and K005 share an identical arrangement of arginine biosynthesis gene while K004 showed a slightly different arrangement by lacking the argF gene. Fosmid Ant4E12, an Antarctic actinobacterial clone, showed a higher synteny with K003/5 than K004 and a similar arginine operon, but in a different genomic context. The Clusters of Orthologous Groups categories assignment of the three fosmids yielded genes that were mostly involved in amino acid and nucleotide metabolism, as well as transport and ribosomal RNA translation, structure and biogenesis. These genomic fragments represent the first sequences to be published from these lineages, providing a cornerstone for future work on this environmentally dominant group.