Dynamic diel proteome and daytime nitrogenase activity supports buoyancy in the cyanobacterium Trichodesmium.

Cyanobacteria of the genus Trichodesmium provide about 80 Tg of fixed nitrogen to the surface ocean per year and contribute to marine biogeochemistry, including the sequestration of carbon dioxide. Trichodesmium fixes nitrogen in the daylight, despite the incompatibility of the nitrogenase enzyme with oxygen produced during photosynthesis. While the mechanisms protecting nitrogenase remain unclear, all proposed strategies require considerable resource investment. Here we identify a crucial benefit of daytime nitrogen fixation in Trichodesmium spp. that may counteract these costs. We analysed diel proteomes of cultured and field populations of Trichodesmium in comparison with the marine diazotroph Crocosphaera watsonii WH8501, which fixes nitrogen at night. Trichodesmium's proteome is extraordinarily dynamic and demonstrates simultaneous photosynthesis and nitrogen fixation, resulting in balanced particulate organic carbon and particulate organic nitrogen production. Unlike Crocosphaera, which produces large quantities of glycogen as an energy store for nitrogenase, proteomic evidence is consistent with the idea that Trichodesmium reduces the need to produce glycogen by supplying energy directly to nitrogenase via soluble ferredoxin charged by the photosynthesis protein PsaC. This minimizes ballast associated with glycogen, reducing cell density and decreasing sinking velocity, thus supporting Trichodesmium's niche as a buoyant, high-light-adapted colony forming cyanobacterium. To occupy its niche of simultaneous nitrogen fixation and photosynthesis, Trichodesmium appears to be a conspicuous consumer of iron, and has therefore developed unique iron-acquisition strategies, including the use of iron-rich dust. Particle capture by buoyant Trichodesmium colonies may increase the residence time and degradation of mineral iron in the euphotic zone. These findings describe how cellular biochemistry defines and reinforces the ecological and biogeochemical function of these keystone marine diazotrophs.

Revised microbial and photochemical triple-oxygen isotope effects improve marine gross oxygen production estimates.

The biogeochemical fluxes that cycle oxygen (O2) play a critical role in regulating Earth's climate and habitability. Triple-oxygen isotope (TOI) compositions of marine dissolved O2 are considered a robust tool for tracing oxygen cycling and quantifying gross photosynthetic O2 production. This method assumes that photosynthesis, microbial respiration, and gas exchange with the atmosphere are the primary influences on dissolved O2 content, and that they have predictable, fixed isotope effects. Despite its widespread use, there are major elements of this approach that remain uncharacterized, including the TOI dynamics of respiration by marine heterotrophic bacteria and abiotic O2 sinks such as the photochemical oxidation of dissolved organic carbon (DOC). Here, we report the TOI fractionation for O2 utilization by two model marine heterotrophs and by abiotic photo-oxidation of representative terrestrial and coastal marine DOC. We demonstrate that TOI slopes associated with these processes span a significant range of the mass-dependent domain (λ = 0.499 to 0.521). A sensitivity analysis reveals that even under moderate productivity and photo-oxidation scenarios, true gross oxygen production may deviate from previous estimates by more than 20% in either direction. By considering a broader suite of oxygen cycle reactions, our findings challenge current gross oxygen production estimates and highlight several paths forward to better understanding the marine oxygen and carbon cycles.

Mechanisms and heterogeneity of in situ mineral processing by the marine nitrogen fixer Trichodesmium revealed by single-colony metaproteomics.

The keystone marine nitrogen fixer Trichodesmium thrives in high-dust environments. While laboratory investigations have observed that Trichodesmium colonies can access the essential nutrient iron from dust particles, less clear are the biochemical strategies underlying particle-colony interactions in nature. Here we demonstrate that Trichodesmium colonies engage with mineral particles in the wild with distinct molecular responses. We encountered particle-laden Trichodesmium colonies at a sampling location in the Southern Caribbean Sea; microscopy and synchrotron-based imaging then demonstrated heterogeneous associations with iron oxide and iron-silicate minerals. Metaproteomic analysis of individual colonies by a new low-biomass approach revealed responses in biogeochemically relevant proteins including photosynthesis proteins and metalloproteins containing iron, nickel, copper, and zinc. The iron-storage protein ferritin was particularly enriched implying accumulation of mineral-derived iron, and multiple iron acquisition pathways including Fe(II), Fe(III), and Fe-siderophore transporters were engaged. While the particles provided key trace metals such as iron and nickel, there was also evidence that Trichodesmium was altering its strategy to confront increased superoxide production and metal exposure. Chemotaxis regulators also responded to mineral presence suggesting involvement in particle entrainment. These molecular responses are fundamental to Trichodesmium's ecological success and global biogeochemical impact, and may contribute to the leaching of particulate trace metals with implications for global iron and carbon cycling.

Spatial Heterogeneity in Particle-Associated, Light-Independent Superoxide Production Within Productive Coastal Waters.

In the marine environment, the reactive oxygen species (ROS) superoxide is produced through a diverse array of light-dependent and light-independent reactions, the latter of which is thought to be primarily controlled by microorganisms. Marine superoxide production influences organic matter remineralization, metal redox cycling, and dissolved oxygen concentrations, yet the relative contributions of different sources to total superoxide production remain poorly constrained. Here we investigate the production, steady-state concentration, and particle-associated nature of light-independent superoxide in productive waters off the northeast coast of North America. We find exceptionally high levels of light-independent superoxide in the marine water column, with concentrations ranging from 10 pM to in excess of 2,000 pM. The highest superoxide concentrations were particle associated in surface seawater and in aphotic seawater collected meters off the seafloor. Filtration of seawater overlying the continental shelf lowered the light-independent, steady-state superoxide concentration by an average of 84%. We identify eukaryotic phytoplankton as the dominant particle-associated source of superoxide to these coastal waters. We contrast these measurements with those collected at an off-shelf station, where superoxide concentrations did not exceed 100 pM, and particles account for an average of 40% of the steady-state superoxide concentration. This study demonstrates the primary role of particles in the production of superoxide in seawater overlying the continental shelf and highlights the importance of light-independent, dissolved-phase reactions in marine ROS production.

Dark biological superoxide production as a significant flux and sink of marine dissolved oxygen.

The balance between sources and sinks of molecular oxygen in the oceans has greatly impacted the composition of Earth's atmosphere since the evolution of oxygenic photosynthesis, thereby exerting key influence on Earth's climate and the redox state of (sub)surface Earth. The canonical source and sink terms of the marine oxygen budget include photosynthesis, respiration, photorespiration, the Mehler reaction, and other smaller terms. However, recent advances in understanding cryptic oxygen cycling, namely the ubiquitous one-electron reduction of O2 to superoxide by microorganisms outside the cell, remains unexplored as a potential player in global oxygen dynamics. Here we show that dark extracellular superoxide production by marine microbes represents a previously unconsidered global oxygen flux and sink comparable in magnitude to other key terms. We estimate that extracellular superoxide production represents a gross oxygen sink comprising about a third of marine gross oxygen production, and a net oxygen sink amounting to 15 to 50% of that. We further demonstrate that this total marine dark extracellular superoxide flux is consistent with concentrations of superoxide in marine environments. These findings underscore prolific marine sources of reactive oxygen species and a complex and dynamic oxygen cycle in which oxygen consumption and corresponding carbon oxidation are not necessarily confined to cell membranes or exclusively related to respiration. This revised model of the marine oxygen cycle will ultimately allow for greater reconciliation among estimates of primary production and respiration and a greater mechanistic understanding of redox cycling in the ocean.