Allen J. Milligan

Climate-driven trends in contemporary ocean productivity

Contributing roughly half of the biosphere’s net primary production (NPP), photosynthesis by oceanic phytoplankton is a vital link in the cycling of carbon between living and inorganic stocks. Each day, more than a hundred million tons of carbon in the form of CO2 are fixed into organic material by these ubiquitous, microscopic plants of the upper ocean, and each day a similar amount of organic carbon is transferred into marine ecosystems by sinking and grazing. The distribution of phytoplankton biomass and NPP is defined by the availability of light and nutrients (nitrogen, phosphate, iron). These growth-limiting factors are in turn regulated by physical processes of ocean circulation, mixed-layer dynamics, upwelling, atmospheric dust deposition, and the solar cycle. Satellite measurements of ocean colour provide a means of quantifying ocean productivity on a global scale and linking its variability to environmental factors. Here we describe global ocean NPP changes detected from space over the past decade. The period is dominated by an initial increase in NPP of 1,930 teragrams of carbon a year (Tg C yr-1), followed by a prolonged decrease averaging 190 Tg C yr-1. These trends are driven by changes occurring in the expansive stratified low-latitude oceans and are tightly coupled to coincident climate variability. This link between the physical environment and ocean biology functions through changes in upper-ocean temperature and stratification, which influence the availability of nutrients for phytoplankton growth. The observed reductions in ocean productivity during the recent post-1999 warming period provide insight on how future climate change can alter marine food webs.

THE ELEMENTAL COMPOSITION OF SOME MARINE PHYTOPLANKTON1

We analyzed the cellular content of C, N, P, S, K, Mg, Ca, Sr, Fe, Mn, Zn, Cu, Co, Cd, and Mo in 15 marine eukaryotic phytoplankton species in culture representing the major marine phyla. All the organisms were grown under identical culture conditions, in a medium designed to allow rapid growth while minimizing precipitation of iron hydroxide. The cellular concentrations of all metals, phosphorus, and sulfur were determined by high‐resolution inductively coupled plasma mass spectrometry (HR‐ICPMS) and those of carbon and nitrogen by a carbon hydrogen nitrogen analyzer. Accuracy of the HR‐ICPMS method was validated by comparison with data obtained with 55Fe radioactive tracer and by a planktonic reference material. The cellular quotas (normalized to P) of trace metals and major cations in the biomass varied by a factor of about 20 among species (except for Cd, which varied over two orders of magnitude) compared with factors of 5 to 10 for major nutrients. Green algae had generally higher C, N, Fe, Zn, and Cu quotas and lower S, K, Ca, Sr, Mn, Co, and Cd quotas than coccolithophores and diatoms. Co and Cd quotas were also lower in diatoms than in coccolithophores. Although trace element quotas are influenced by a variety of growth conditions, a comparison of our results with published data suggests that the measured compositions reflect chiefly the intrinsic (i.e. genetically encoded) trace element physiology of the individual species. Published field data on the composition of the planktonic biomass fall within the range of laboratory values and are generally close to the approximate extended Redfield formula given by the average stoichiometry of our model species (excluding the hard parts): While clearly this elemental stoichiometry varies between species and, potentially, in response to changes in the chemistry of seawater, it provides a basis for examining how phytoplankton influence the relative distributions of the ensemble of major and trace elements in the ocean.

Evolved physiological responses of phytoplankton to their integrated growth environment

Phytoplankton growth and productivity relies on light, multiple nutrients and temperature. These combined factors constitute the ‘integrated growth environment’. Since their emergence in the Archaean ocean, phytoplankton have experienced dramatic shifts in their integrated growth environment and, in response, evolved diverse mechanisms to maximize growth by optimizing the allocation of photosynthetic resources (ATP and NADPH) among all cellular processes. Consequently, co-limitation has become an omnipresent condition in the global ocean. Here we focus on evolved phytoplankton populations of the contemporary ocean and the varied energetic pathways they employ to solve the optimization problem of resource supply and demand. Central to this discussion is the allocation of reductant formed through photosynthesis, which we propose has the following three primary fates: carbon fixation, direct use and ATP generation. Investment of reductant among these three sinks is tied to cell cycle events, differentially influenced by specific forms of nutrient stress, and a strong determinant of relationships between light-harvesting (pigment), photosynthetic electron transport and carbon fixation. Global implications of optimization are illustrated by deconvolving trends in the 10-year global satellite chlorophyll record into contributions from biomass and physiology, thereby providing a unique perspective on the dynamic nature of surface phytoplankton populations and their link to climate.

8.5 – Marine Bioinorganic Chemistry: The Role of Trace Metals in the Oceanic Cycles of Major Nutrients

The bulk of living biomass is chiefly made up of only a dozen ‘major’ elements whose proportions vary within a relatively narrow range in most organisms. A number of trace elements, particularly first row transition metals are also ‘essential’ for the growth of organisms. We begin this chapter by discussing what we know of the concentrations of trace elements in marine microorganisms and of the relevant mechanisms and kinetics of trace-metal uptake. We then review the biochemical role of trace elements in the marine cycles of carbon, nitrogen, phosphorus, and silicon. Using this information, we examine the evidence, emanating from both laboratory cultures and field measurements, relevant to the mechanisms and the extent of control by trace metals of marine biogeochemical cycles. Before concluding with a wistful glimpse of the future of marine bioinorganic chemistry we discuss briefly some paleoceanographic aspects of this new field: how the chemistry of the planet ‘Earth’ – particularly the concentrations of trace elements in the oceans – has evolved since its origin, chiefly as a result of biological processes and how the evolution of life has, in turn, been affected by the availability of essential trace elements.

The Role of the C4 Pathway in Carbon Accumulation and Fixation in a Marine Diatom

The role of a C4 pathway in photosynthetic carbon fixation by marine diatoms is presently debated. Previous labeling studies have shown the transfer of photosynthetically fixed carbon through a C4 pathway and recent genomic data provide evidence for the existence of key enzymes involved in C4 metabolism. Nonetheless, the importance of the C4 pathway in photosynthesis has been questioned and this pathway is seen as redundant to the known CO2 concentrating mechanism of diatoms. Here we show that the inhibition of phosphoenolpyruvate carboxylase (PEPCase) by 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate resulted in a more than 90% decrease in whole cell photosynthesis in Thalassiosira weissflogii cells acclimated to low CO2 (10 μm), but had little effect on photosynthesis in the C3 marine Chlorophyte, Chlamydomonas sp. In 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate-treated T. weissflogii cells, elevated CO2 (150 μm) or low O2 (80–180 μm) restored photosynthesis to the control rate linking PEPCase inhibition with CO2 supply in this diatom. In C4 organic carbon-inorganic carbon competition experiments, the 12C-labeled C4 products of PEPCase, oxaloacetic acid and its reduced form malic acid suppressed the fixation of 14C-labeled inorganic carbon by 40% to 50%, but had no effect on O2 evolution in photosynthesizing diatoms. Oxaloacetic acid-dependent O2 evolution in T. weissflogii was twice as high in cells acclimated to 10 μm rather than 22 μm CO2, indicating that the use of C4 compounds for photosynthesis is regulated over the range of CO2 concentrations observed in marine surface waters. Short-term 14C uptake (silicone oil centrifugation) and CO2 release (membrane inlet mass spectrometry) experiments that employed a protein denaturing cell extraction solution containing the PEPCKase inhibitor mercaptopicolinic acid revealed that much of the carbon taken up by diatoms during photosynthesis is stored as organic carbon before being fixed in the Calvin cycle, as expected if the C4 pathway functions as a CO2 concentrating mechanism. Together these results demonstrate that the C4 pathway is important in carbon accumulation and photosynthetic carbon fixation in diatoms at low (atmospheric) CO2.

Photophysiological Expressions of Iron Stress in Phytoplankton

Iron is essential for all life, but it is particularly important to photoautotrophs because of the many iron-dependent electron transport components in photosynthetic membranes. Since the proliferation of oxygenic photosynthesis in the Archean ocean, iron has been a scarce commodity, and it is now recognized as a limiting resource for phytoplankton over broad expanses of the open ocean and even in some coastal/continental shelf waters. Iron stress does not impair photochemical or carbon fixation efficiencies, and in this respect it resembles the highly tuned photosynthetic systems of steady-state macronutrient-limited phytoplankton. However, iron stress does present unique photophysiological challenges, and phytoplankton have responded to these challenges through major architectural changes in photosynthetic membranes. These evolved responses include overexpression of photosynthetic pigments and iron-economic pathways for ATP synthesis, and they result in diagnostic fluorescence properties that allow a broad appraisal of iron stress in the field and even the detection of iron stress from space.

Acquisition of inorganic carbon by the marine diatom Thalassiosira weissflogii

Recent data on the physiology of inorganic carbon acquisition by the model marine diatom Thalassiosira weissflogii (Grunow) demonstrate the importance of the catalytic equilibration of HCO3-and CO2by carbonic anhydrases located in the periplasm and in the cytoplasm. These enzymes can use Zn, Co or Cd as their metal centre, and their activity increases at low ambient CO2. The silica frustule provides buffering for extracellular CA activity, The transmembrane transport of CO2 may occur by passive diffusion. Under CO2 limitation, the cytoplasmic HCO3-is used to form malate and oxaloacetic acid via phosphoenolpyruvate carboxylase. It appears that subsequent decarboxylation of these compounds in the chloroplast regenerates CO2 near the site of Rubisco, and thus provides the organism with an effective unicellular C4 photosynthetic pathway. These results, together with other published data, bring up two major questions regarding inorganic carbon acquisition in diatoms: What is the major species of inorganic carbon (CO2 or HCO3-) transported across the membrane under natural conditions? And what is the form of carbon (inorganic or organic) accumulated by the cells?

Surplus Photosynthetic Antennae Complexes Underlie Diagnostics of Iron Limitation in a Cyanobacterium

Chlorophyll fluorescence from phytoplankton provides a tool to assess iron limitation in the oceans, but the physiological mechanism underlying the fluorescence response is not understood. We examined fluorescence properties of the model cyanobacterium Synechocystis PCC6803 and a ΔisiA knock-out mutant of the same species grown under three culture conditions which simulate nutrient conditions found in the open ocean: (1) nitrate and iron replete, (2) limiting-iron and high-nitrate, representative of natural high-nitrate, low-chlorophyll regions, and (3) iron and nitrogen co-limiting. We show that low variable fluorescence, a key diagnostic of iron limitation, results from synthesis of antennae complexes far in excess of what can be accommodated by the iron-restricted pool of photosynthetic reaction centers. Under iron and nitrogen co-limiting conditions, there are no excess antennae complexes and variable fluorescence is high. These results help to explain the well-established fluorescence characteristics of phytoplankton in high-nutrient, low-chlorophyll ocean regions, while also accounting for the lack of these properties in low-iron, low-nitrogen regions. Importantly, our results complete the link between unique molecular consequences of iron stress in phytoplankton and global detection of iron stress in natural populations from space.

LINKING TIME-DEPENDENT CARBON-FIXATION EFFICIENCIES IN DUNALIELLA TERTIOLECTA (CHLOROPHYCEAE) TO UNDERLYING METABOLIC PATHWAYS1

The chl‐specific short‐term 14C‐based production (Pb) measurement is a widely used tool to understand phytoplankton responses to environmental stresses. However, among the metabolic consequences of these stresses is variability in lifetimes of newly fixed carbon that cause Pb to range between chl‐specific net primary production (NPP*) and chl‐specific gross photosynthetic electron flow that is available for carbon reduction () depending on growth rate. To investigate the basis for this discrepancy, photosynthate utilization was characterized in Dunaliella tertiolecta Butcher grown at three different growth rates in N‐limited chemostats. Pb was measured throughout a 2 min to 24 h time course and showed clear growth‐rate‐dependent differences in lifetimes of newly fixed carbon. 14C pulse‐chase experiments revealed differences in patterns of carbon utilization between growth rates. At high growth rate, the majority of 14C was initially fixed into polysaccharide and lipid, but the relative contribution of each labeled biochemical pool to the total label changed over 24 h. In fast‐growing cells, labeled polysaccharides decreased 50%, while labeled lipids increased over the first 4 h. At low growth rate, 14C was initially incorporated primarily into protein, but the contribution of labeled protein to the total label increased over the next 24 h. Together, time‐resolved measurements of Pb and cellular NAD and NADP content suggest an enhanced role for alternative dissipation pathways at very low growth rate. Findings of this study contribute to an integrated understanding of growth‐rate‐dependent shifts in metabolic processes from photosynthesis to net growth.

A common partitioning strategy for photosynthetic products in evolutionarily distinct phytoplankton species

· We compare the nutrient-dependent photosynthetic efficiencies of the chlorophyte, Dunaliella tertiolecta, with those of the marine diatom, Thalassiosira weissflogii. Despite considerable evolutionary and physiological differences, these two species appear to use nearly identical growth strategies under a wide range of nutrient limitation. · Using a variety of physiological measurements, we find that, for both species and across all growth rates, 75% of the gross photosynthetic electron flow is invested in carbon fixation and only 30% is retained as net carbon accumulation. A majority of gross photosynthesis (70%) is ultimately used as reductant for biosynthetic pathways and for the generation of ATP. · In both species, newly formed carbon products exhibit much shorter half-lives at slow growth rates than at fast growth rates. We show that this growth rate dependence is a result of increased polysaccharide storage during the S phase of the cell cycle. · We present a model of carbon utilization that incorporates this growth rate-dependent carbon allocation and accurately captures (r(2) = 0.94) the observed time-resolved carbon retention. Together, our findings suggest a common photosynthetic optimization strategy in evolutionarily distinct phytoplankton species and contribute towards a systems-level understanding of carbon flow in photoautotrophs.