Effect of iron on the growth of Phaeodactylum tricornutum via photosynthesis

Iron is a limiting factor that controls the phytoplankton biomass in the modern ocean, and iron fertilization of the ocean could lead to blooms dominated by diatoms. Thus, iron plays an important role in controlling the distribution of diatoms. In this study, we measured the growth rate and photosynthetic activity of the model diatom Phaeodactylum tricornutum cultured under different iron concentrations and found that it grew more rapidly and had a much higher photosynthetic efficiency under higher iron concentrations. In order to explore the unique mechanism of the response of diatoms to iron, a proteomic analysis was carried out, and the results indicated that iron promotes the Calvin cycle of P. tricornutum. Diatoms can tolerate the pressure of iron limitation by replacing iron‐rich proteins with flavodoxin, and so on. Moreover, we found that the photosystem I (PSI) activity of iron‐limited algae that were treated by N’,N’,N’,N’‐tetramethyl‐p‐phenylenediamine (TMPD) was increased significantly. As TMPD plays the role of a cytochrome b₆/f complex that transfers electrons from photosystem II to PSI, the cytochrome b₆/f complex is the key to photosynthesis regulation. Iron could influence the growth of P. tricornutum by regulating its biosynthesis. All of the results suggest that iron might affect the growth of diatoms through the Calvin cycle and the cytochrome b₆/f complex.     

Limitations on microalgal growth at very low photon fluence rates: the role of energy slippage

The lower limits of photosynthetically useable radiation at which growth and photosynthesis can occur establish the lower boundaries for the extent of photolithotrophy in the biosphere. Photolithotrophic growth denotes the capacity to grow with photons as the sole energy input. Slippage in terms of photosynthetic energy conversion implies a less than theoretical stoichiometry of energy-transduction process(es) such as the dissipation of intermediates of O₂ evolution and of ATP synthesis (H⁺/e⁻ and H⁺/ATP ratios). Slippage is particularly important in limiting the growth of photolithotrophic organisms at very low photon fluence rates. We found that Dunaliella tertiolecta and Phaeodactylum tricornutum avoid such reductions in photon use efficiency by increasing the size and number of their photosynthetic units, respectively, and by altering Q_A reduction kinetics on the reducing side of PS II. P. tricornutum is also less susceptible to slippage in terms of the breakdown of intermediates in its O₂ evolution pathway than D. tertiolecta. Minimizing H⁺ leakage through the CF₀–CF₁ ATP synthetase (and other H⁺ porters) is also discussed briefly. In combination, strategies employed by P.␣tricornutum effectively allow it to grow and photosynthesize at lower rates of energy input than D. tertiolecta, consistent with our observations. Differences in the responses of the photosynthetic apparatus of these two marine microalgae are mechanistic and probably representative of evolutionary divergences associated with strategies for dealing with environmental perturbations.     

STATE TRANSITIONS AND NONPHOTOCHEMICAL QUENCHING DURING A NUTRIENT‐INDUCED FLUORESCENCE TRANSIENT IN PHOSPHORUS‐STARVED DUNALIELLA TERTIOLECTA1

Assessments of nutrient‐limitation in microalgae using chl a fluorescence have revealed that nitrogen and phosphorus depletion can be detected as a change in chl a fluorescence signal when nutrient‐starved algae are resupplied with the limiting nutrient. This photokinetic phenomenon is known as a nutrient‐induced fluorescence transient, or NIFT. Cultures of the unicellular marine chlorophyte Dunaliella tertiolecta Butcher were grown under phosphate starvation to investigate the photophysiological mechanism behind the NIFT response. A combination of low temperature (77 K) fluorescence, photosynthetic inhibitors, and nonphotochemical quenching analyses were used to determine that the NIFT response is associated with changes in energy distribution between PSI and PSII and light‐stress‐induced nonphotochemical quenching (NPQ). Previous studies point to state transitions as the likely mechanism behind the NIFT response; however, our results show that state transitions are not solely responsible for this phenomenon. This study shows that an interaction of at least two physiological processes is involved in the rapid quenching of chl a fluorescence observed in P‐starved D. tertiolecta: (1) state transitions to provide the nutrient‐deficient cell with metabolic energy for inorganic phosphate (P_i)‐uptake and (2) energy‐dependent quenching to allow the nutrient‐stressed cell to avoid photodamage from excess light energy during nutrient uptake.     

Protein turnover in relation to maintenance metabolism at low photon flux in two marine microalgae

Acclimation to very low photon fluxes involves adjusting a suite of physiological characteristics that collectively elicit a physiological response. Facilitating such changes is pro‐tein turnover. Dunaliella tertiolecta (Butcher) and Phaeodactylum tricornutum (Bohlin) were grown in turbidostats at a range of photon fluxes between 2 and 300 µmol photons m^?2 s^?1. The kinetics of pulse‐chase labelling of the protein with ³H showed that (1) two protein pools were present, one of which turned‐over rapidly (hours), and a second which turned over more slowly (days); and (2) protein turnover rates were slower in P. tricornutum than in D. tertiolecta. Phaeodactylum tricornutum had a lower maintenance coefficient for protein turnover than D. tertiolecta, and correspondingly a smaller proportion of its respiratory demands (30%) were associated with protein turnover than in D. tertiolecta (36%). There appears to be a correlation between lower metabolic activity, requiring lower protein concentrations, and an associated decreased cost of maintenance processes in P. tricornutum compared to D. tertiolecta. Differences between protein turnover rates and maintenance metabolic costs may be one of the photo‐acclimation strategies that determine which photon niches microalgae can successfully exploit.     

The effect of cadmium and lead on the growth of two species of marine phytoplankton with particular reference to the development of tolerance

Decreases in salinity (<10%) increased the growth rates ofPhaeodactylum tricornutum and Dunaliella tertiolecta. Increasinglevels of cadmium (1–50 ppm (mg 1^–1)) reduced thegrowth rates of both species. 100 ppm cadmium was lethal toD. tertiolecta but not to P. tricornutum. Lead (1 –4 ppm)initially increased the growth rate of D. tertiolecta but thencaused all but the 1 ppm culture to die. Lead (1–4 ppm)caused a decrease in growth rate of P. tricornutum. After exposureto 1 ppm cadmium, cultures of D. tertiolecta showed an increasedtolerance to levels of cadmium, and a changed response to levelsof lead. Exposure of P. tricornutum to either cadmium or lead,or exposure of D. tertiolecta to lead caused no change in responseto either metal.     

RESPONSE OF THE PHOTOSYNTHETIC APPARATUS OF PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE) TO NITRATE,PHOSPHATE, OR IRON STARVATION1

The effects of nitrate, phosphate, and iron starvation and resupply on photosynthetic pigments, selected photosynthetic proteins, and photosystem II (PSII) photochemistry were examined in the diatom Phaeodactylum tricornutum Bohlin (CCMP 1327). Although cell chlorophyll a (chl a) content decreased in nutrient-starved cells, the ratios of light-harvesting accessory pigments (chl c and fucoxanthin) to chl a were unaffected by nutrient starvation. The chl a-specific light absorpition coefficient (a*) and the functional absorption cross-section of PSII (σ) increased during nutrient starvation, consistent with reduction of intracellular self-shading (i.e. a reduction of the “package effect”) as cells became chlorotic. The light-harvesting complex proteins remained a constant proportion of total cell protein during nutrient starvation, indicating that chlorosis mirrored a general reduction in cell protein content. The ratio of the xanthophylls cycle pigments diatoxanthin and diadinoxanthin to chl a increased during nutrient starvation. These pigments are thought to play a photo-protective role by increasing dissipation of excitation energy in the pigment bed upstream from the reaction centers. Despite the increase in diatoxanthin and diadinoxanthin, the efficiency of PSII photochemistry, as measured by the ration of variable to maximum fluorescence (F_v/F_m) of dark-adapted cells, declined markedly under nitrate and iron starvation and moderately under phosphate starvation. Parallel to changes in F_v/F_m were decreases in abundance of the reaction center protein D1 consistent with damage of PSII reaction centers in nutrient-starved cells. The relative abundance of the carboxylating enzyme, ribulose bisphosphate carboxylase/oxygenase (RUBISCO), decreased in response to nitrate and iron starvation but not phosphate starvation. Most marked was the decline in the abundance of the small subunit of RUBISCO in nitrate-starved cells. The changes in pigment content and fluorescence characteristics were typically reversed within 24 h of resupply of the limiting nutrient.     

Comparison of population growth and photosynthetic apparatus changes in response to different nutrient status in a diatom and a coccolithophore

In many marine ecosystems, diatoms dominate in nutrient‐rich coastal waters while coccolithiophores are found offshore in areas where nutrients may be limiting. In lab‐controlled batch cultures, mixed‐species competition between the diatom Phaeodactylum tricornutum and the coccolithophore Emiliana huxleyi and the response of each species were examined under nitrate (N) and phosphate (P) starvation. Based on the logistic growth model and the Lotka–Volterra competition model, E. huxleyi showed higher competitive abilities than P. tricornutum under N and P starvation. For both species, cell growth was more inhibited by P starvation, while photosynthetic functions (chl a fluorescence parameters) and cellular constituents (pigments) were impaired by N starvation. The decline of photosynthetic functions occurred later in E. huxleyi (day 12) than in P. tricornutum (day 9); this time difference was associated with greater damage of the photosynthetic apparatus in P. tricornutum compared with E. huxleyi. Xanthophyll cycle pigment accumulation and the transformation from diadinoxanthin to diatoxanthin was more active in E. huxleyi than P. tricornutum, under similar N and P starvation. We concluded that E. huxleyi and P. tricornutum have different mechanisms to allocate resources and energy under nutrient starvation. It appears that E. huxleyi has a more economic strategy to adapt to nutrient depleted environments than P. tricornutum. These findings provided additional evidence explaining how N versus P limitation differentially support diatom and coccolithophore blooms in natural environments.     

Iron-induced changes in light harvesting and photochemical energy conversion processes in eukaryotic marine algae

The role of iron in regulating light harvesting and photochemical energy conversion processes was examined in the marine unicellular chlorophyte Dunaliella tertiolecta and the marine diatom Phaeodactylum tricornutum. In both species, iron limitation led to a reduction in cellular chlorophyll concentrations, but an increase in the in vivo, chlorophyll-specific, optical absorption cross-sections. Moreover, the absorption cross-section of photosystem II, a measure of the photon target area of the traps, was higher in iron-limited cells and decreased rapidly following iron addition. Iron-limited cells exhibited reduced variable/maximum fluorescence ratios and a reduced fluorescence per unit absorption at all wave-lengths between 400 and 575 nm. Following iron addition, variable/maximum fluorescence ratios increased rapidly, reaching 90% of the maximum within 18 to 25 h. Thus, although more light was absorbed per unit of chlorophyll, iron limitation reduced the transfer efficiency of excitation energy in photosystem II. The half-time for the oxidation of primary electron acceptor of photosystem II, calculated from the kinetics of decay of variable maximum fluorescence, increased 2-fold under iron limitation. Quantitative analysis of western blots revealed that cytochrome f and subunit IV (the plastoquinone-docking protein) of the cytochrome b₆/f complex were also significantly reduced by lack of iron; recovery from iron limitation was completely inhibited by either cycloheximide or chloramphenicol. The recovery of maximum photosynthetic energy conversion efficiency occurs in three stages: (a) a rapid (3-5 h) increase in electron transfer rates on the acceptor side of photosystem II correlated with de novo synthesis of the cytochrome b₆/f complex; (b) an increase (10-15 h) in the quantum efficiency correlated with an increase in D1 accumulation; and (c) a slow (>18 h) increase in chlorophyll levels accompanied by an increase in the efficiency of energy transfer from the light-harvesting chlorophyll proteins to the reaction centers.     

AMMONIUM INDUCED PHOTOSYNTHETIC SUPPRESSION IN AMMONIUM LIMITED DUNALIELLA TERTIOLECTA (CHLOROPHYTA)1

Dunaliella tertiolecta (Butcher) was grown in chemostat culture over a wide range of ammonium limited growth rates. The addition of ammonium caused a rapid temporary suppression of photosynthetic carbon fixation. The magnitude of ammonium induced photosynthetic suppression increased with the severity of ammonium limitation. Cells growing at rates greater than ca. 80%μ_max exhibited no photosynthetic suppression in response to additions of the limiting nutrient. The duration of photosynthetic suppression was related to the concentration of added ammonium. Immediately following the suppression, photosynthesis was enhanced with respect to the controls. The eventual degree of enhancement increased with the concentration of added ammonium. Steady-state cellular chlorophyll concentrations, photosynthetic rates, and assimilation numbers are reported.     

干热河谷车桑子光合生理特性对氮磷添加的响应

氮磷养分是限制干热河谷植物生长的重要元素,不同土壤上植物受到的养分限制类型不同。光合作用作为植物生长发育的基础,不同土壤上氮磷养分添加对干热河谷植物光合生理特征的影响还没有报道。因此,以干热河谷优势植物——车桑子为研究对象,在元谋县不同海拔处采集土壤,设置加氮（+N）、加磷（+P）、氮磷同时添加（+NP）和不添加（CK）四个处理,研究车桑子光合响应曲线、叶绿素含量和叶绿素荧光特性对氮、磷添加的响应规律,并探讨光合响应特征与车桑子生长的关系：研究结果显示：1）不同海拔土壤上,车桑子光合生理特性对氮磷添加具有不同的响应。在低海拔燥红土上,氮添加处理（+N和+NP）提高了车桑子净光合速率、叶绿素含量和PSII活性;中海拔紫色土上,+NP促进了车桑子光合速率和叶绿素含量的提高;高海拔黄棕壤上,+N处理降低了车桑子净光合速率和PSII活性,而磷添加处理（+P和+NP）提高了车桑子净光合速率。2）车桑子光合特性对养分添加的响应取决于土壤的养分限制类型,限制性养分添加可以提高车桑子的净光合速率。3）燥红土上+P以及黄棕壤上+N对PSⅡ最大光化学效率（Fv/Fm）的降低更大程度上归于可变荧光Fv的减少而不是最小荧光F₀的增加,可减少养分限制对光系统II造成的伤害。4）三种土壤类型上车桑子的叶绿素含量和组成差异极显著,相比于燥红土和紫色土,黄棕壤上车桑子的叶绿素含量显著更高,而叶绿素a/b显著更低。综上,本研究结果表明,车桑子光合能力受到氮和磷的共同调节,不同土壤上光合生理特性的响应可增强植物对限制性养分的适应性,影响植物生长发育。     

INTERACTIONS AMONG PHOSPHATE UPTAKE,PHOTOSYNTHESIS, AND CHLOROPHYLL FLUORESCENCE IN NUTRIENT‐LIMITED CULTURES OF THE CHLOROPHYTE MICROALGA DUNALIELLA TERTIOLECTA1

Phosphate‐limited and phosphate‐sufficient continuous cultures of the marine chlorophyte microalga Dunaliella tertiolecta Butcher were examined for their responses to the addition of phosphate. Phosphate‐limited cultures showed a marked quenching of chl fluorescence following a pulse of phosphate. This response was absent from cells growing under phosphate‐sufficient conditions. Both the extent of fluorescence quenching (where present) and the initial rate of change in quenching were dependent on the concentration of phosphate added to cell suspensions and on the degree of limitation (growth rate in continuous culture). The addition of phosphate also brought about a transient decrease in photosynthetic oxygen evolution and a stimulation in respiration, which were relaxed as the added phosphate was depleted from the external medium. The applicability of using nutrient‐induced fluorescence transients as a tool to identify the nutrient status of phytoplankton populations is discussed.     

VARIABILITY IN CELL SIZE,NUTRIENT DEPLETION,AND GROWTH RATES OF THE SOUTHERN OCEAN DIATOM FRAGILARIOPSIS KERGUELENSIS (BACILLARIOPHYCEAE) AFTER PROLONGED IRON LIMITATION1

Diatoms are the main primary producers in the Southern Ocean, governing the major nutrient cycles. Fragilariopsis kerguelensis (O’Meara) Hust. is the most abundant diatom species in the Southern Ocean and its paleo‐oceanographic record is frequently used to reconstruct the past position and nutrient characteristics of the Antarctic polar front. Here we report on the responses of F. kerguelensis on prolonged exposure to a range of iron concentrations, allowing a characterization of morphological and nutrient‐depletion changes in relation to iron status. Under iron limitation, F. kerguelensis grew slower, cells became smaller, chains became shorter, and the nutrient‐depletion ratios changed. Prolonged exposure to iron limitation caused F. kerguelensis to decrease its surface area and volume 2‐fold, and to increase its surface‐to‐volume ratio by 25%. With the decrease in growth rates, silicon (Si) and phosphorus (P) depletion per cell remained fairly constant, but when normalized per surface area (Si) or per cell volume (P), depletion increased. In contrast, nitrogen (N) depletion per cell decreased significantly together with the decrease in growth rates but was constant when normalized per cell volume. The different response in Si, P, and N depletion resulted in changes in the nutrient‐depletion ratios, most notably in the Si:N ratio, which significantly increased, and in the N:P ratio, which significantly decreased with decreasing growth rates. It is concluded that under iron limitation, variation in cell size and/or nutrient depletion ultimately can cause changes in oceanic biogeochemical nutrient cycles. It enables the use of cell size of F. kerguelensis as a paleo‐oceanographic proxy.     

FIRST INDUCED PLASTID GENOME MUTATIONS IN AN ALGA WITH SECONDARY PLASTIDS: psbA MUTATIONS IN THE DIATOM PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE) REVEAL CONSEQUENCES ON THE REGULATION OF PHOTOSYNTHESIS1

Diatoms play a crucial role in the biochemistry and ecology of most aquatic ecosystems, especially because of their high photosynthetic productivity. They often have to cope with a fluctuating light climate and a punctuated exposure to excess light, which can be harmful for photosynthesis. To gain insight into the regulation of photosynthesis in diatoms, we generated and studied mutants of the diatom Phaeodactylum tricornutum Bohlin carrying functionally altered versions of the plastidic psbA gene encoding the D1 protein of the PSII reaction center (PSII RC). All analyzed mutants feature an amino acid substitution in the vicinity of the Q_B‐binding pocket of D1. We characterized the photosynthetic capacity of the mutants in comparison to wildtype cells, focusing on the way they regulate their photochemistry as a function of light intensity. The results show that the mutations resulted in constitutive changes of PSII electron transport rates. The extent of the impairment varies between mutants depending on the proximity of the mutation to the Q_B‐binding pocket and/or to the nonheme iron within the PSII RC. The effects of the mutations described here for P. tricornutum are similar to effects in cyanobacteria and green microalgae, emphasizing the conservation of the D1 protein structure among photosynthetic organisms of different evolutionary origins.     

Interaction Effects of Ambient UV Radiation and Nutrient Limitation on the Toxic Cyanobacterium <Emphasis Type="Italic">Nodularia spumigena</Emphasis>

Nodularia spumigena is one of the dominating species during the extensive cyanobacterial blooms in the Baltic Sea. The blooms coincide with strong light, stable stratification, low ratios of dissolved inorganic nitrogen, and dissolved inorganic phosphorus. The ability of nitrogen fixation, a high tolerance to phosphorus starvation, and different photo-protective strategies (production of mycosporine-like amino acids, MAAs) may give N. spumigena a competitive advantage over other phytoplankton during the blooms. To elucidate the interactive effects of ambient UV radiation and nutrient limitation on the performance of N. spumigena, an outdoor experiment was designed. Two radiation treatments photosynthetic active radiation (PAR) and PAR +UV-A + UV-B (PAB) and three nutrient treatments were established: nutrient replete (NP), nitrogen limited (−N), and phosphorus limited (−P). Variables measured were specific growth rate, heterocyst frequency, cell volume, cell concentrations of MAAs, photosynthetic pigments, particulate carbon (POC), particulate nitrogen (PON), and particulate phosphorus (POP). Ratios of particulate organic matter were calculated: POC/PON, POC/POP, and PON/POP. There was no interactive effect between radiation and nutrient limitation on the specific growth rate of N. spumigena, but there was an overall effect of phosphorus limitation on the variables measured. Interaction effects were observed for some variables; cell size (larger cells in −P PAB compared to other treatments) and the carotenoid canthaxanthin (highest concentration in −N PAR). In addition, significantly less POC and PON (mol cell⁻¹) were found in −P PAR compared to −P PAB, and the opposite radiation effect was observed in −N. Our study shows that despite interactive effects on some of the variables studied, N. spumigena tolerate high ambient UVR also under nutrient limiting conditions and maintain positive growth rate even under severe phosphorus limitation.     

Nutrient and iron limitation to <Emphasis Type="Italic">Ulva</Emphasis> blooms in a eutrophic coastal lagoon (Sacca di Goro,Italy)

Growth patterns and bloom formation of the green seaweed Ulva rigida were analysed in the eutrophic Sacca di Goro lagoon (Po River Delta, Italy). Variations of standing biomasses and elemental composition of Ulva were analysed through an annual cycle with respect to nitrogen, phosphorus and iron. Growth rates, nutrient and iron uptake and nitrate storage by macroalgal thalli were also assessed with field experiments during the formation of a spring bloom. The control of Ulva growth and the bloom formation depended on multiple factors, especially on nitrogen availability and iron deficiency. In the nitrate rich waters of the Sacca di Goro lagoon, nitrate accumulation in Ulva thalli was inversely related with Fe uptake, indicating an influence of Fe limitation on N acquisition. Since length and magnitude of nitrate luxury uptake are inversely related to the size of the intracellular nitrate pools, in nitrate rich waters the fast growing Ulva may face risk of N-limitation not only when exposed to low N concentrations or at high biomass levels, but also when exposed to pulsed dissolved nitrate concentrations at low iron availability. The potential Fe limitation could be affected by processes controlled by geochemical reactions and by macroalgal growth and decomposition. Both Fe oxidation during the active macroalgal growth and the formation of insoluble FeS and FeS₂ during bloom collapse can result in a drastic decrease of soluble iron. Thus, a potential limitation of Fe to macroalgae can occur, determining positive feedbacks and potentially controlling the extent of bloom development and persistence.     

Light-, nitrogen-, and phosphorus-limited growth of Phaeodactylum tricornutum Bohlin strain TFX-1: Chemical composition,carbon partitioning,and the diel periodicity of physiological processes

Phaeodactylum tricomutum Bohlin (strain TFX-1) was grown under light-, nitrogen-, and phosphorus-limited conditions in continuous or semicontinuous cultures under a 12L-12D light regime. The C, N, and P contents were determined at each steady state, as was the partitioning of cellular organic carbon into protein, lipids, polysaccharides, and metabolic intermediates. All determinations were made at the beginning and again at the end of the light period. The rates of nutrient assimilation and of synthesis of biochemical constituents during the light and dark periods were calculated from the above data, and the periodicities of these processes characterized. The elemental composition of the cells was different under each limitation. In particular, phosphorus limitation severely restricted the ability of the cell to store nitrogen in non-protein forms. Biochemical composition and the diel periodicity of cellular processes also differed between limitations. Nutrient uptake was most strongly periodic under light limitation. Protein synthesis showed increased periodicity under nitrogen limitation, relative to the other limitations, while the periodicity of lipid synthesis was reduced under phosphate limitation. Polysaccharide was synthesized at high rates during the light period and consumed in the dark under all limitations.     

Plasticity in photosynthetic response to nutrient supply of seedlings from a mixed conifer‐angiosperm forest

Abstract We measured the plasticity of the response of photosynthesis to nutrient supply in seedlings of the dominant four conifer and broadleaved angiosperm tree species from an indigenous forest in South‐westland, New Zealand. We hypothesized that the response of conifers to differing nutrient supply would be less than the response for the angiosperms because of greater adaptation to low fertility conditions. In Prumnopitys ferruginea (D. Don) de Laub. the maximum velocity of electron transport, J_max, doubled with a 10‐fold increase in concentration of nitrogen supply. In Dacrydium cupressinum Lamb. the maximum velocity of carboxylation, V_cmax, doubled with a 10‐fold increase in phosphorus supply. In contrast, photosynthetic capacity for the angiosperm species Weinmannia racemosa L.f. was affected only by the interaction of nitrogen and phosphorus and photosynthetic capacity of Metrosideros umbellata Cav. was not affected by nutrient supply. The response of the conifers to increasing availability of nutrient suggests greater plasticity in photosynthetic capacity, a characteristic not generally associated with adaptation to soil infertility, thus invalidating our hypothesis. Our data suggest that photosynthetic response to nutrient supply cannot be broadly generalized between the two functional groups.