Phosphorus and nitrogen starvation reveal life-cycle specific responses in the metabolome of Emiliania huxleyi (Haptophyta)

The coccolithophore Emiliania huxleyi is a microalga with biogeochemical and biotechnological relevance, due to its high abundance in the ocean and its ability to form intricate calcium carbonate structures. Depletion of macronutrients in oceanic waters is very common and will likely enhance with advancing climate change. We present the first comprehensive metabolome study analyzing the effect of phosphorus (P) and nitrogen (N) starvation on the diploid and haploid life-cycle stage, applying various metabolome analysis methods to gain new insights in intracellular mechanisms to cope with nutrient starvation. P-starvation led to an accumulation of many generic and especially N-rich metabolites, including lipids, osmolytes, and pigments. This suggests that P-starvation primarily arrests cell-cycling due to lacking P for nucleic acid synthesis, but that enzymatic functionality is widely preserved. Also, the de-epoxidation ratio of the xanthophyll cycle was upregulated in the diploid stage under P-starvation, indicating increased nonphotochemical quenching, a response typically observed under high light stress. In contrast, N-starvation resulted in a decrease of most central metabolites, also P-containing ones, especially in the diploid stage, indicating that most enzymatic functionality ceased. The two investigated nutrient starvation conditions caused significantly different responses, contrary to previous assumptions derived from transcriptomic studies. Data highlight that instantaneous biochemical flux is a more dominant driver of the metabolome than the transcriptomically rearranged pathway patterns. Due to the fundamental nature of the observed responses it may be speculated that microalgae with similar nutrient requirements can cope better with P-starvation than with N-starvation

Temperature Modulates Coccolithophorid Sensitivity of Growth, Photosynthesis and Calcification to Increasing Seawater pCO2

Increasing atmospheric CO2 concentrations are expected to impact pelagic ecosystem functioning in the near future by driving ocean warming and acidification. While numerous studies have investigated impacts of rising temperature and seawater acidification on planktonic organisms separately, little is presently known on their combined effects. To test for possible synergistic effects we exposed two coccolithophore species, Emiliania huxleyi and Gephyrocapsa oceanica, to a CO2 gradient ranging from ,0.5–250 mmol kg21 (i.e. ,20–6000 matm pCO2) at three different temperatures (i.e. 10, 15, 20uC for E. huxleyi and 15, 20, 25uC for G. oceanica). Both species showed CO2-dependent optimum-curve responses for growth, photosynthesis and calcification rates at all temperatures. Increased temperature generally enhanced growth and production rates and modified sensitivities of metabolic processes to increasing CO2. CO2 optimum concentrations for growth, calcification, and organic carbon fixation rates were only marginally influenced from low to intermediate temperatures. However, there was a clear optimum shift towards higher CO2 concentrations from intermediate to high temperatures in both species. Our results demonstrate that the CO2 concentration where optimum growth, calcification and carbon fixation rates occur is modulated by temperature. Thus, the response of a coccolithophore strain to ocean acidification at a given temperature can be negative, neutral or positive depending on that strain’s temperature optimum. This emphasizes that the cellular responses of coccolithophores to ocean acidification can only be judged accurately when interpreted in the proper eco-physiological context of a given strain or species. Addressing the synergistic effects of changing carbonate chemistry and temperature is an essential step when assessing the success of coccolithophores in the future ocean

MRI of intact plants

Nuclear magnetic resonance imaging (MRI) is a non-destructive and non-invasive technique that can be used to acquire two- or even three-dimensional images of intact plants. The information within the images can be manipulated and used to study the dynamics of plant water relations and water transport in the stem, e.g., as a function of environmental (stress) conditions. Non-spatially resolved portable NMR is becoming available to study leaf water content and distribution of water in different (sub-cellular) compartments. These parameters directly relate to stomatal water conductance, CO2 uptake, and photosynthesis. MRI applied on plants is not a straight forward extension of the methods discussed for (bio)medical MRI. This educational review explains the basic physical principles of plant MRI, with a focus on the spatial resolution, factors that determine the spatial resolution, and its unique information for applications in plant water relations that directly relate to plant photosynthetic activity

Essential omega‐3 fatty acids are depleted in sea ice and pelagic algae of the Central Arctic Ocean

Microalgae are the main source of the omega‐3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), essential for the healthy development of most marine and terrestrial fauna including humans. Inverse correlations of algal EPA and DHA proportions (% of total fatty acids) with temperature have led to suggestions of a warming‐induced decline in the global production of these biomolecules and an enhanced importance of high latitude organisms for their provision. The cold Arctic Ocean is a potential hotspot of EPA and DHA production, but consequences of global warming are unknown. Here, we combine a full‐seasonal EPA and DHA dataset from the Central Arctic Ocean (CAO), with results from 13 previous field studies and 32 cultured algal strains to examine five potential climate change effects; ice algae loss, community shifts, increase in light, nutrients, and temperature. The algal EPA and DHA proportions were lower in the ice‐covered CAO than in warmer peripheral shelf seas, which indicates that the paradigm of an inverse correlation of EPA and DHA proportions with temperature may not hold in the Arctic. We found no systematic differences in the summed EPA and DHA proportions of sea ice versus pelagic algae, and in diatoms versus non‐diatoms. Overall, the algal EPA and DHA proportions varied up to four‐fold seasonally and 10‐fold regionally, pointing to strong light and nutrient limitations in the CAO. Where these limitations ease in a warming Arctic, EPA and DHA proportions are likely to increase alongside increasing primary production, with nutritional benefits for a non‐ice‐associated food web

Effects of CO2 and their modulation by light in the life-cycle stages of the coccolithophore Emiliania huxleyi

The effects of ocean acidification on the life-cycle stages of the coccolithophore Emiliania huxleyi and their modulation by light were examined. Calcifying diploid and noncalcifying haploid cells (Roscoff culture collection strains 1216 and 1217) were acclimated to present-day and elevated CO2 partial pressures (PCO2; 38.5 vs. 101.3 Pa, i.e., 380 vs. 1000 µatm) under low and high light (50 vs. 300 µmol photons m-2 s-1). Growth rates as well as cellular quotas and production rates of C and N were measured. Sources of inorganic C for biomass buildup were determined using a 14C disequilibrium assay. Photosynthetic O2 evolution was measured as a function of dissolved inorganic C and light by means of membrane-inlet mass spectrometry. The diploid stage responded to elevated PCO2 by shunting resources from the production of particulate inorganic C toward organic C yet keeping the production of total particulate C constant. As the effect of ocean acidification was stronger under low light, the diploid stage might be less affected by increased acidity when energy availability is high. The haploid stage maintained elemental composition and production rates under elevated PCO2. Although both life-cycle stages involve different ways of dealing with elevated PCO2, the responses were generally modulated by energy availability, being typically most pronounced under low light. Additionally, PCO2 responses resembled those induced by high irradiances, indicating that ocean acidification affects the interplay between energy-generating processes (photosynthetic light reactions) and processes competing for energy (biomass buildup and calcification). A conceptual model is put forward explaining why the magnitude of single responses is determined by energy availability

Physiological effects of ocean acidification on the coccolithophore Emiliania huxleyi and their modulation by light

To investigate the modulation of OA-responses by light intensity, cells of E. huxleyi strain RCC 1216 were acclimated to ambient and high pCO2 (380 vs. 1000 µatm) under limiting and saturating light intensities (50 vs. 300 µmol photons m-2 s-1). Growth rates and cellular quotas of POC and PIC were measured. Photosynthetic O2 evolution was measured as a function of light and of [DIC]

Characterization of the life-cycle stages of the coccolithophore Emiliania huxleyi and their responses to Ocean Acidification

Anthropogenic carbon dioxide emissions cause a chemical phenomenon known as Ocean Acidification (OA). The associated changes in seawater chemistry are believed to have significant impact especially on coccolithophores, unicellular calcifying primary producers that take an outstanding role in the regulation of the marine carbon pumps. This thesis investigated the calcifying diploid and the non-calcifying haploid life-cycle stages of the globally dominant coccolithophore Emiliania huxleyi, and their responses to OA. Emphasis was put on investigating the role of energy-availability (i.e., irradiance) in the manifestation of OA-responses. A suite of methods was applied to resolve the effects on the phenomenological level (growth, elemental quotas and production), the physiological level (photosynthesis, carbon acquisition) and the level of gene expression (transcriptomics). In publication I, haploid and diploid cells were compared using microarray-based transcriptome profiling to assess stage-specific gene expression. The study identified genes related to distinct cell-biological traits, such as calcification in the diplont as well as flagellae and lipid respiration in the haplont. It further revealed that the diploid stage needs to make more regulatory efforts to epigenetically administrate its double amount of DNA, and therefore strongly controls its gene expression on the basis of transcription. The haplont in turn, possessing only a single sized genome, does not require these administrative efforts and seems to drive a more unrestricted gene expression. The proteome is apparently regulated on the basis of rapid turnover, i.e., post-translational. The haploid and diploid genomes may therefore be regarded as cellular ‘operating systems’ that streamline the life-cycle stages to occupy distinct ecological niches. Publication II investigated the responses of the life-cycle stages to OA under limiting and saturating light intensities. Growth rates as well as quotas and production rates of carbon (C) and nitrogen (N) were measured. In addition, inorganic C acquisition and photosynthesis were determined with a 14C-tracer technique and mass spectrometrybased gas-flux measurements. Under OA, the diploid stage shunted resources from calcification towards biomass production, yet keeping the production of total particulate carbon constant. In the haploid stage, elemental composition and production rates were more or less unaffected although major physiological acclimations were evident, pointing towards efforts to maintain homeostasis. Apparently, both life-cycle stages pursue distinct strategies to deal with OA. As a general pattern, OA-responses were strongly modulated by energy availability and typically most pronounced under low light. A concept explaining the energy-dependence of responses was proposed. In publication III, microarray-based transcriptome profiling was used to screen for cellular processes that underlie the observed phenomenological and physiological responses observed in the life-cycle stages (publication II). In the diplont, the increased biomass production under OA seems to be caused by production of glycoconjugates and lipids. 8 The lowered calcification may be attributed to impaired signal-transduction and iontransport mechanisms. The haplont utilized genes and metabolic pathways distinct from the diploid stage, reflecting the stage-specific usage of certain portions of the genome. With respect to functionality and energy-dependence, however, the transcriptomic OAresponses resembled those of the diplont. In both stages, signal transduction and ionhomeostasis were equally OA-sensitive under all light intensities. The effects on carbon metabolism and photophysiology, however, were clearly modulated by light availability. These interactive effects can be explained with the influence of both OA and light on the cellular ‘redox hub’, a major sensory system controlling the network of metabolic sources and sinks of reductive energy. In the general discussion, the newly gained views on the life-cycle stages are synthesized and biogeochemical implications of light-dependent OA-effects on coccolithophore calcification are considered. Furthermore, emerging physiological response patterns are identified to develop unifying concepts that can explain the energy-dependence of physiological effects. Finally, the critical role of redox regulation in the responses to changing environmental parameters is argued and research perspectives are given how to further resolve effects of the changing environment on marine phytoplankton