The genus Nitrospira is the most widespread group of nitrite-oxidizing bacteria and thrives in diverse natural and engineered ecosystems. Nitrospira marina Nb-295
was isolated from the ocean over 30 ...years ago; however, its genome has not yet been analyzed. Here, we investigated the metabolic potential of N. marina based on its complete genome sequence and performed physiological experiments to test genome-derived hypotheses. Our data confirm that N. marina benefits from additions of undefined organic carbon substrates, has adaptations to resist oxidative, osmotic, and UV light-induced stress and low dissolved pCO
, and requires exogenous vitamin B
. In addition, N. marina is able to grow chemoorganotrophically on formate, and is thus not an obligate chemolithoautotroph. We further investigated the proteomic response of N. marina to low (∼5.6 µM) O
concentrations. The abundance of a potentially more efficient CO
-fixing pyruvate:ferredoxin oxidoreductase (POR) complex and a high-affinity cbb
-type terminal oxidase increased under O
limitation, suggesting a role in sustaining nitrite oxidation-driven autotrophy. This putatively more O
-sensitive POR complex might be protected from oxidative damage by Cu/Zn-binding superoxide dismutase, which also increased in abundance under low O
conditions. Furthermore, the upregulation of proteins involved in alternative energy metabolisms, including Group 3b NiFe hydrogenase and formate dehydrogenase, indicate a high metabolic versatility to survive conditions unfavorable for aerobic nitrite oxidation. In summary, the genome and proteome of the first marine Nitrospira isolate identifies adaptations to life in the oxic ocean and provides insights into the metabolic diversity and niche differentiation of NOB in marine environments.
Thaumarchaeota are among the most abundant microbial cells in the ocean, but difficulty in cultivating marine Thaumarchaeota has hindered investigation into the physiological and evolutionary basis ...of their success. We report here a closed genome assembled from a highly enriched culture of the ammonia-oxidizing pelagic thaumarchaeon CN25, originating from the open ocean. The CN25 genome exhibits strong evidence of genome streamlining, including a 1.23-Mbp genome, a high coding density, and a low number of paralogous genes. Proteomic analysis recovered nearly 70% of the predicted proteins encoded by the genome, demonstrating that a high fraction of the genome is translated. In contrast to other minimal marine microbes that acquire, rather than synthesize, cofactors, CN25 encodes and expresses near-complete biosynthetic pathways for multiple vitamins. Metagenomic fragment recruitment indicated the presence of DNA sequences >90% identical to the CN25 genome throughout the oligotrophic ocean. We propose the provisional name "Candidatus Nitrosopelagicus brevis" str. CN25 for this minimalist marine thaumarchaeon and suggest it as a potential model system for understanding archaeal adaptation to the open ocean.
Marine primary productivity is strongly influenced by the scarcity of required nutrients, yet our understanding of these nutrient limitations is informed by experimental observations with sparse ...geographical coverage and methodological limitations. We developed a quantitative proteomic method to directly assess nutrient stress in high-light ecotypes of the abundant cyanobacterium Prochlorococcus across a meridional transect in the central Pacific Ocean. Multiple peptide biomarkers detected widespread and overlapping regions of nutritional stress for nitrogen and phosphorus in the North Pacific Subtropical Gyre and iron in the equatorial Pacific. Quantitative protein analyses demonstrated simultaneous stress for these nutrients at biome interfaces. This application of proteomic biomarkers to diagnose ocean metabolism demonstrated Prochlorococcus actively and simultaneously deploying multiple biochemical strategies for low-nutrient conditions in the oceans.
The marine nitrogen fixing microorganisms (diazotrophs) are a major source of nitrogen to open ocean ecosystems and are predicted to be limited by iron in most marine environments. Here we use global ...and targeted proteomic analyses on a key unicellular marine diazotroph Crocosphaera watsonii to reveal large scale diel changes in its proteome, including substantial variations in concentrations of iron metalloproteins involved in nitrogen fixation and photosynthesis, as well as nocturnal flavodoxin production. The daily synthesis and degradation of enzymes in coordination with their utilization results in a lowered cellular metalloenzyme inventory that requires ~40% less iron than if these enzymes were maintained throughout the diel cycle. This strategy is energetically expensive, but appears to serve as an important adaptation for confronting the iron scarcity of the open oceans. A global numerical model of ocean circulation, biogeochemistry and ecosystems suggests that Crocosphaera's ability to reduce its iron-metalloenzyme inventory provides two advantages: It allows Crocosphaera to inhabit regions lower in iron and allows the same iron supply to support higher Crocosphaera biomass and nitrogen fixation than if they did not have this reduced iron requirement.
We prepared highly oriented, multi-lamellar stacks of human red blood cell (RBC) membranes applied on silicon wafers. RBC ghosts were prepared by hemolysis and applied onto functionalized silicon ...chips and annealed into multi-lamellar RBC membranes. High resolution X-ray diffraction was used to determine the molecular structure of the stacked membranes. We present direct experimental evidence that these RBC membranes consist of nanometer sized domains of integral coiled-coil peptides, as well as liquid ordered (l
) and liquid disordered (l
) lipids. Lamellar spacings, membrane and hydration water layer thicknesses, areas per lipid tail and domain sizes were determined. The common drug aspirin was added to the RBC membranes and found to interact with RBC membranes and preferably partition in the head group region of the l
domain leading to a fluidification of the membranes, i.e., a thinning of the bilayers and an increase in lipid tail spacing. Our results further support current models of RBC membranes as patchy structures and provide unprecedented structural details of the molecular organization in the different domains.
Scarce dissolved surface ocean concentrations of the essential algal micronutrient zinc suggest that Zn may influence the growth of phytoplankton such as diatoms, which are major contributors to ...marine primary productivity. However, the specific mechanisms by which diatoms acclimate to Zn deficiency are poorly understood. Using global proteomic analysis, we identified two proteins (ZCRP-A/B, Zn/Co Responsive Protein A/B) among four diatom species that became abundant under Zn/Co limitation. Characterization using reverse genetic techniques and homology data suggests putative Zn/Co chaperone and membrane-bound transport complex component roles for ZCRP-A (a COG0523 domain protein) and ZCRP-B, respectively. Metaproteomic detection of ZCRPs along a Pacific Ocean transect revealed increased abundances at the surface (<200 m) where dZn and dCo were scarcest, implying Zn nutritional stress in marine algae is more prevalent than previously recognized. These results demonstrate multiple adaptive responses to Zn scarcity in marine diatoms that are deployed in low Zn regions of the Pacific Ocean.
The diffusion of membrane receptors is central to many biological processes, such as signal transduction, molecule translocation, and ion transport, among others; consequently, several advanced ...fluorescence microscopy techniques have been developed to measure membrane receptor mobility within live cells. The membrane-anchored receptor cluster of differentiation 14 (CD14) and the transmembrane toll-like receptor 2 (TLR2) are important receptors in the plasma membrane of macrophages that activate the intracellular signaling cascade in response to pathogenic stimuli. The aim of the present work was to compare the diffusion coefficients of CD14 and TLR2 on the apical and basal membranes of macrophages using two fluorescence-based methods: raster image correlation spectroscopy (RICS) and single particle tracking (SPT). In the basal membrane, the diffusion coefficients obtained from SPT and RICS were found to be comparable and revealed significantly faster diffusion of CD14 compared with TLR2. In addition, RICS showed that the diffusion of both receptors was significantly faster in the apical membrane than in the basal membrane, suggesting diffusion hindrance by the adhesion of the cells to the substrate. This finding highlights the importance of selecting the appropriate membrane (i.e., basal or apical) and corresponding method when measuring receptor diffusion in live cells. Accurately knowing the diffusion coefficient of two macrophage receptors involved in the response to pathogen insults will facilitate the study of changes that occur in signaling in these cells as a result of aging and disease.
Coastal Antarctic marine ecosystems are significant in carbon cycling because of their intense seasonal phytoplankton blooms. Southern Ocean algae are primarily limited by light and iron (Fe) and can ...be co-limited by cobalamin (vitamin B
). Micronutrient limitation controls productivity and shapes the composition of blooms which are typically dominated by either diatoms or the haptophyte
. However, the vitamin requirements and ecophysiology of the keystone species
remain poorly characterized. Using cultures, physiological analysis, and comparative omics, we examined the response of
to a matrix of Fe-B
conditions. We show that
is not auxotrophic for B
, as previously suggested, and identify mechanisms underlying its B
response in cultures of predominantly solitary and colonial cells. A combination of proteomics and proteogenomics reveals a B
-independent methionine synthase fusion protein (MetE-fusion) that is expressed under vitamin limitation and interreplaced with the B
-dependent isoform under replete conditions. Database searches return homologues of the MetE-fusion protein in multiple
species and in a wide range of marine microbes, including other photosynthetic eukaryotes with polymorphic life cycles as well as bacterioplankton. Furthermore, we find MetE-fusion homologues expressed in metaproteomic and metatranscriptomic field samples in polar and more geographically widespread regions. As climate change impacts micronutrient availability in the coastal Southern Ocean, our finding that
has a flexible B
metabolism has implications for its relative fitness compared to B
-auxotrophic diatoms and for the detection of B
-stress in a more diverse set of marine microbes.
Summary
Diatoms are a highly successful group of photosynthetic protists that often thrive under adverse environmental conditions. Members of the genus Pseudo‐nitzschia are ecologically important ...diatoms which are able to subsist during periods of chronic iron limitation and form dense blooms following iron fertilization events. The cellular strategies within diatoms that orchestrate these physiological responses to variable iron concentrations remain largely uncharacterized. Using a combined transcriptomic and proteomic approach, we explore the exceptional ability of a diatom isolated from the iron‐limited Northeast Pacific Ocean to reorganize its intracellular processes as a function of iron. We compared the molecular responses of Pseudo‐nitzschia granii observed under iron‐replete and iron‐limited growth conditions to those of other model diatoms. Iron‐coordinated molecular responses demonstrated some agreement between gene expression and protein abundance, including iron‐starvation‐induced‐proteins, a putative iron transport system and components of photosynthesis and the Calvin cycle. Pseudo‐nitzschia granii distinctly differentially expresses genes encoding proteins involved in iron‐independent photosynthetic electron transport, urea acquisition and vitamin synthesis. We show that P. granii is unique among studied diatoms in its physiology stemming from distinct cellular responses, which may underlie its ability to subsist in low iron regions and rapidly bloom to outcompete other diatom taxa following iron enrichment.
Bioactive trace metals are critical micronutrients for marine
microorganisms due to their role in mediating biological redox reactions,
and complex biogeochemical processes control their ...distributions.
Hydrothermal vents may represent an important source of metals to
microorganisms, especially those inhabiting low-iron waters, such as in the
southwest Pacific Ocean. Previous measurements of primordial 3He
indicate a significant hydrothermal source originating in the northeastern (NE)
Lau Basin, with the plume advecting into the southwest Pacific Ocean at
1500–2000 m depth (Lupton et
al., 2004). Studies investigating the long-range transport of trace metals
associated with such dispersing plumes are rare, and the biogeochemical
impacts on local microbial physiology have not yet been described. Here we
quantified dissolved metals and assessed microbial metaproteomes across a
transect spanning the tropical and equatorial Pacific with a focus on the
hydrothermally active NE Lau Basin and report elevated iron and manganese
concentrations across 441 km of the southwest Pacific. The most intense
signal was detected near the Mangatolo Triple Junction (MTJ) and Northeast
Lau Spreading Center (NELSC), in close proximity to the previously reported
3He signature. Protein content in distal-plume-influenced seawater,
which was high in metals, was overall similar to background locations,
though key prokaryotic proteins involved in metal and organic uptake,
protein degradation, and chemoautotrophy were abundant compared to deep
waters outside of the distal plume. Our results demonstrate that trace
metals derived from the NE Lau Basin are transported over appreciable
distances into the southwest Pacific Ocean and that bioactive chemical
resources released from submarine vent systems are utilized by surrounding
deep-sea microbes, influencing both their physiology and their contributions
to ocean biogeochemical cycling.
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