Snow Alga Chlamydomonas Nivalis Research Articles

Quantitative proteomic comparison of salt stress in Chlamydomonas reinhardtii and the snow alga Chlamydomonas nivalis reveals mechanisms for salt-triggered fatty acid accumulation via reallocation of carbon resources

BackgroundChlamydomonas reinhardtii is a model green alga strain for molecular studies; its fully sequenced genome has enabled omic-based analyses that have been applied to better understand its metabolic responses to stress. Here, we characterised physiological and proteomic changes between a low-starch C. reinhardtii strain and the snow alga Chlamydomonas nivalis, to reveal insights into their contrasting responses to salinity stress.ResultsEach strain was grown in conditions tailored to their growth requirements to encourage maximal fatty acid (as a proxy measure of lipid) production, with internal controls to allow comparison points. In 0.2 M NaCl, C. nivalis accumulates carbohydrates up to 10.4% DCW at 80 h, and fatty acids up to 52.0% dry cell weight (DCW) over 12 days, however, C. reinhardtii does not show fatty acid accumulation over time, and shows limited carbohydrate accumulation up to 5.5% DCW. Analysis of the C. nivalis fatty acid profiles showed that salt stress improved the biofuel qualities over time. Photosynthesis and respiration rates are reduced in C. reinhardtii relative to C. nivalis in response to 0.2 M NaCl. De novo sequencing and homology matching was used in conjunction with iTRAQ-based quantitative analysis to identify and relatively quantify proteomic alterations in cells exposed to salt stress. There were abundance differences in proteins associated with stress, photosynthesis, carbohydrate and lipid metabolism proteins. In terms of lipid synthesis, salt stress induced an increase in dihydrolipoyl dehydrogenase in C. nivalis (1.1-fold change), whilst levels in C. reinhardtii remained unaffected; this enzyme is involved in acetyl CoA production and has been linked to TAG accumulation in microalgae. In salt-stressed C. nivalis there were decreases in the abundance of UDP-sulfoquinovose (− 1.77-fold change), which is involved in sulfoquinovosyl diacylglycerol metabolism, and in citrate synthase (− 2.7-fold change), also involved in the TCA cycle. Decreases in these enzymes have been shown to lead to increased TAG production as fatty acid biosynthesis is favoured. Data are available via ProteomeXchange with identifier PXD018148.ConclusionsThese differences in protein abundance have given greater understanding of the mechanism by which salt stress promotes fatty acid accumulation in the un-sequenced microalga C. nivalis as it switches to a non-growth state, whereas C. reinhardtii does not have this response.

Cold Adaptation Mechanisms of a Snow Alga Chlamydomonas nivalis During Temperature Fluctuations.

Cold environments, such as glaciers and alpine regions, constitute unique habitats for organisms living on Earth. In these harsh ecosystems, snow algae survive, florish, and even become primary producers for microbial communities. How the snow algae maintain physiological activity during violent ambient temperature changes remains unsolved. To explore the cold adaptation mechanisms of the unicellular snow alga Chlamydomonas nivalis, we compared its physiological responses to a model organism from the same genus, Chlamydomonas reinhardtii. When both cell types were exposed to a shift from 22°C to 4°C, C. nivalis exhibited an apparent advantage in cold tolerance over C. reinhardtii, as C. nivalis had both a higher growth rate and photosynthetic efficiency. To determine the cold tolerance mechanisms of C. nivalis, RNA sequencing was used to compare transcriptomes of both species after 1 h of cold treatment, mimicking temperature fluctuations in the polar region. Differential expression analysis showed that C. nivalis had fewer transcriptomic changes and was more stable during rapid temperature decrease relative to C. reinhardtii, especially for the expression of photosynthesis related genes. Additionally, we found that transcription in C. nivalis was precisely regulated by the cold response network, consisting of at least 12 transcription factors and 3 RNA-binding proteins. Moreover, genes participating in nitrogen metabolism, the pentose phosphate pathway, and polysaccharide biosynthesis were upregulated, indicating that increasing resource assimilation and remodeling of metabolisms were critical for cold adaptation in C. nivalis. Furthermore, we identified horizontally transferred genes differentially expressed in C. nivalis, which are critical for cold adaptation in other psychrophiles. Our results reveal that C. nivalis adapts rapid temperature decrease by efficiently regulating transcription of specific genes to optimize resource assimilation and metabolic pathways, providing critical insights into how snow algae survive and propagate in cold environments.

Low-Temperature Adaptation of the Snow Alga Chlamydomonas nivalis Is Associated With the Photosynthetic System Regulatory Process.

The alga Chlamydomonas nivalis thrives in polar snow fields and on high-altitude mountain tops, and contributes significantly on primary production in the polar regions, however, the mechanisms underlying this adaptation to low temperatures are unknown. Here, we compared the growth, photosynthetic activity, membrane lipid peroxidation, and antioxidant activity of C. nivalis with those of the model alga C. reinhardtii, under grow temperature and low temperatures. C. nivalis maintained its photosynthetic activity in these conditions by reducing the light-harvesting ability of photosystem II and enhancing the cyclic electron transfer around photosystem I, both of which limited damage to the photosystem from excess light energy and resulted in ATP production, supporting cellular growth and other physiological processes. Furthermore, the increased cyclic electron transfer rate, carotenoid content, and antioxidant enzyme activities jointly regulated the reactive oxygen species levels in C. nivalis, enabling recovery from excess excitation energy and reduced photooxidative damage to the cell. Therefore, we propose a model in which adaptive mechanisms related to photosynthetic regulation promote the survival and even blooming of C. nivalis under polar environment, suggesting that C. nivalis can provide organic carbon sources as an important primary producer for other surrounding life in the polar regions for maintaining ecosystem.

Global Metabolic Regulation of the Snow Alga Chlamydomonas nivalis in Response to Nitrate or Phosphate Deprivation by a Metabolome Profile Analysis.

In the present work, Chlamydomonas nivalis, a model species of snow algae, was used to illustrate the metabolic regulation mechanism of microalgae under nutrient deprivation stress. The seed culture was inoculated into the medium without nitrate or phosphate to reveal the cell responses by a metabolome profile analysis using gas chromatography time-of-flight mass spectrometry (GC/TOF-MS). One hundred and seventy-one of the identified metabolites clustered into five groups by the orthogonal partial least squares discriminant analysis (OPLS-DA) model. Among them, thirty of the metabolites in the nitrate-deprived group and thirty-nine of the metabolites in the phosphate-deprived group were selected and identified as “responding biomarkers” by this metabolomic approach. A significant change in the abundance of biomarkers indicated that the enhanced biosynthesis of carbohydrates and fatty acids coupled with the decreased biosynthesis of amino acids, N-compounds and organic acids in all the stress groups. The up- or down-regulation of these biomarkers in the metabolic network provides new insights into the global metabolic regulation and internal relationships within amino acid and fatty acid synthesis, glycolysis, the tricarboxylic acid cycle (TCA) and the Calvin cycle in the snow alga under nitrate or phosphate deprivation stress.

135 Measured glass transition temperatures within cells following slow cooling: Implications for cryopreservation and freeze drying

Following slow rates of cooling, at which cellular dehydration occurs, it is assumed that the intracellular environment vitrifies. However, the glass transition (Tg) temperature has not previously been determined for any cell type during cooling. Using differential scanning calorimetry (DSC), we have measured intracellularTg, in a range of cell types (bacteria, algae, yeasts) when cooled slowly in the presence of extracellular ice, where no cryoprotectants were added. Under the experimental conditions used, any intracellular freezing would have been detected by the release of latent heat associated with the change in entropy that accompanies the phase transition from liquid water to ice in the cell during cooling, or the reverse during warming; none was seen. Vitrification was detected by the variation in heat flow associated with the change in heat capacity, together with the absence of any release of latent heat. No vitrification signal was seen in control runs (medium without cells). The signal detected in the sample runs was the glass transition temperature of the maximally freeze-concentrated intracellular compartment (conventionally designated Tg’). The observed values of Tg’ ranged from just below −10 °C in thermophilic bacteria and the ice-nucleating bacterium Pseudomonas syringae, to around −25 °C in three species, including the snow alga Chlamydomonas nivalis. These temperatures are well above typical environmental temperatures for high latitudes in winter, indicating that intracellular vitrification must be a process of widespread ecological relevance for free-living single cells. These findings have a number of implications for cryopreservation: (a) During slow cooling, vitrification will occur at a high sub zero temperature and the cell will be osmotically unresponsive during subsequent cooling. On warming the vitrified cells will be exposed to an extracellular hypertonic stress before the cell interior become liquid. This set of physical events are not described in any current model of freezing injury, it is assumed that the cell is osmotically reactive at all temperatures down to the Tg of the extracellular solution. (b) Vitrification would be expected to stop any damaging intracellular events, but damage to the plasmalemma may still occur at the outer surface by direct exposure to hypertonic solutions. (c) Freeze drying of cells will always occur at a temperature below the intracellular vitrification temperature Source of funding: INRA, CEPIA Department, UMR 782 GMPA, F 78850 Thiverval Grignon, France. Conflict of interest: None declared. fonseca@grignon.inra.fr

Lipidomic profiling reveals lipid regulation in the snow alga Chlamydomonas nivalis in response to nitrate or phosphate deprivation

The snow alga Chlamydomonas nivalis in the exponential phase was subjected to nitrate or phosphate deprivation for 0–72h to study its stress responses in lipid profiles analyzed by UPLC/Q-TOF-MS (ultra performance liquid chromatography/quadrupole-time of flight-mass spectrometry). Three clusters were distinguished as the control, nitrate-deprived and phosphate-deprived groups in OPLS-DA (orthogonal projection on latent structure discriminant analysis) score plots based on their lipidome data. Altogether, the lipidomic approach identified twenty-two ions in nitrate-deprived group including nine l, 2-diacylglyceryl-3-O-4′-(N, N, N-trimethyl)-homoserine (DGTSs), one phosphatidylethanolamine (PE), two monogalactosyldiacylglycerols (MGDGs), four digalactosyldiacylglycerols (DGDGs), three phosphatidylglycerols (PGs), two sulfoquinovosyl-diacylglycerols (SQDGs) and one phosphatidylinositiol (PI), and nineteen ions in phosphate-deprived group including four DGTSs, one PE, one MGDG, seven DGDGs, three PGs, two SQDGs and one PG as “differentiating lipid biomarkers”. Moreover, the common and specific biomarkers were found in the two nutrient deprived groups by SUS (shared and unique structure) plot. Biomarkers-based z-score plot and heat map further showed how lipid biomarker expressions deviate from the control. The up- or down-regulation of these lipid biomarkers provided new insights into the lipid metabolism of the snow alga in response to nitrate or phosphate deprivation stress condition.

LC–MS/APCI identification of glucoside esters and diesters of astaxanthin from the snow alga Chlamydomonas nivalis including their optical stereoisomers

HPLC methods (LC-MS/APCI and chiral HPLC) were used for the identification of astaxanthin derivatives from the red snow alga Chlamydomonas nivalis collected in Austrian Alps, Slovak High Tatra Mountains and Bulgarian Pirin. We observed a striking difference in the composition of astaxanthin optical isomers in C. nivalis collected in geographically distinct regions. Furthermore, algae from the Pirin Mountains differed in the dominance of astaxanthin diglucoside diesters, suggesting an alternative strategy to enhance cell viability at low temperatures.

Fatty Acids Profiling and Biomarker Identification in Snow Alga Chlamydomonas Nivalis by NaCl Stress Using GC/MS and Multivariate Statistical Analysis

The snow alga Chlamydomonas nivalis is a model species of microalgae for the investigation of cell response mechanism and adaptation ability in natural habitats in polar regions and similar extreme environments. The alteration of fatty acids in cellular lipids is known to play a vital role for cell survival and reproduction under various stress conditions. In the present work, an integrated approach of gas chromatography/mass spectrometry (GC/MS) coupled with multivariate statistical analysis was developed to investigate the fatty acid profiles and identify the biomarkers in response to NaCl stress. The data of fatty acid profiles between the control and NaCl-stress group was classified by orthogonal projection on latent structure discriminant analysis (OPLS-DA) and hierarchical cluster analysis (HCA); six of fatty acids (C16:0, C16:3, C18:0, C18:1, C18:2, and C18:3) were identified as biomarkers. These biomarkers showed a regulatory role by decreasing the degree of lipid unsaturation (DLU), providing an expected function in reducing membrane fluidity and permeability for enhancing the tolerance to higher salinity. This is the first report to demonstrate the fatty acid biomarkers in microalgae as the physiological regulators corresponding to the response and adaptation to NaCl stress based on an integrated approach at the lipidomic level.

Regulation of lipid metabolism in the snow alga Chlamydomonas nivalis in response to NaCl stress: An integrated analysis by cytomic and lipidomic approaches

The cultures of the snow alga Chlamydomonas nivalis in the exponential phage were stressed by NaCl (up to 1.5%) for 0∼48h, followed by Nile Red staining-based cytomic analysis (flow cytometry and confocal laser scanning microscopy). The fluorescent intensities of total lipids, and neutral and polar lipids increased to the maximum within 7h in the NaCl stressed cells with the highest increase in total lipids by 2-fold (0.75%-NaCl for 7h), the highest increase in neutral lipids by 68-fold (1%-NaCl for 7h) and the highest increase in polar lipids by 10-fold (1.25%-NaCl for 5h), respectively. Seven types and 22 kinds of polar lipid molecules were selected and identified as biomarkers by UPLC/Q-TOF-MS-based lipidomic analysis, which demonstrated differences in total lipids between the stress group (0.75%-NaCl for 7h) and the control. The biological roles of the biomarkers in the alga under NaCl stress were discussed. The integrated approach based on “omics” technologies developed in the present work is validated as a powerful tool to successfully reveal the regulation of lipid metabolism in microalgae in response to stress stimulation.

Lipidomic profiling and discovery of lipid biomarkers in snow alga Chlamydomonas nivalis under salt stress

AbstractThe snow alga Chlamydomonas nivalis is a typical microalgal species that can adapt and resist to natural habitats in the polar region and similar extreme environments. In response to various stress conditions, the change of lipid profile in cell membrane systems is known to play a vital role in cell survival and reproduction. In the present work, to elucidate the involvement of intracellular lipids in the molecular mechanism of cell response and adaptation to salt stress, C. nivalis was cultured and treated with different sodium chloride (NaCl) concentrations in different time courses. The ultra performance liquid chromatography/quadrupole‐TOF‐MS (UPLC/Q‐TOF‐MS)‐based approach was developed for lipidomic profiling followed by multivariate statistical analysis including partial least squares discriminant analysis and orthogonal projection on latent structure discriminant analysis for data classification and potential biomarkers selection. Seven types and 35 kinds of polar lipid molecules were selected and identified as biomarkers, including monogalactosyldiacylglycerol, digalactosyldiacylglycerol, l, 2‐diacylglyceryl‐3‐O‐4′‐(N,N,N‐trimethyl)‐homoserine, phosphatidylglycerol, sulfoquinovosyl‐diacylglycerol, phosphatidylethanolamine, and phosphatidylinositiol. Their functions in relation to the regulation of cell membrane stability, signal transduction and photosynthesis efficiency under salt stress are also discussed in this paper. This is the first study using UPLC/Q‐TOF‐MS‐based lipidomic profiling with multivariate statistical analysis for lipid biomarkers discovery from microalgae in response to stress conditions.

Retinal very long-chain PUFAs: new insights from studies on ELOVL4 protein

Compared with other mammalian tissues, retina is highly enriched in PUFA. Long-chain PUFA (LC-PUFA; C18-C24) are essential FAs that are enriched in the retina and are necessary for maintenance of normal retinal development and function. The retina, brain, and sperm also contain very LC-PUFA (VLC-PUFA; >C24). Although VLC-PUFA were discovered more than two decades ago, very little is known about their biosynthesis and functional roles in the retina. This is due mainly to intrinsic difficulties associated with working on these unusually long polyunsaturated hydrocarbon chains and their existence in small amounts. Recent studies on the FA elongase elongation of very long chain fatty acids-4 (ELOVL4) protein, however, suggest that VLC-PUFA probably play some uniquely important roles in the retina as well as the other tissues. Mutations in the ELOVL4 gene are found in patients with autosomal dominant Stargardt disease. Here, we review the recent literature on VLC-PUFA with special emphasis on the elongases responsible for their synthesis. We focus on a novel elongase, ELOVL4, involved in the synthesis of VLC-PUFA, and the importance of these FAs in maintaining the structural and functional integrity of retinal photoreceptors.

Effects of realistically simulated, elevated UV irradiation on photosynthesis and pigment composition of the alpine snow alga Chlamydomonas nivalis and the arctic soil alga Tetracystis sp. (Chlorophyceae)

An indoor sun simulator was used to provide elevated UV-B radiation (280–315 nm) in combination with realistic ratios to PAR (400–700 nm) and UV-A radiation (315–400 nm) in order to test the physiological response of a soil- and snow microalga during a three-day stress scenario, which may occasionally occur in their respective arctic and alpine habitats. Chlamydomonas nivalis and Tetracystis sp. are initial colonizers of harsh habitats like summer snow fields and bare arctic soils. The two species were chosen because of their role as primary successors in places where life is generally limited by extreme climatic and nutritional conditions. The influence of the increased UV-B irradiation (1.43 W m –2 ; control: 0.52 W m –2 ) on photosynthesis and pigment composition was measured. Both species survived this incubation without any morphological signs of damage, but oxygen production was reduced by 20–56%. Under control conditions, the amount of chlorophylls (Chls) and carotenoids (Cars) per dry mass increased after three days due to optimal light conditions. After the same period, the treated samples of the soil alga Tetracystis sp. showed a smaller increase in Chls and primary Cars than the control. However, the production of extraplastidal, secondary Cars was induced. On the contrary, the snow alga C. nivalis already had high amounts of secondary Cars before the experiment, and after exposure, all pigment classes increased more compared to control conditions. The results show that these microalgae can tolerate short episodes of enhanced UV-B radiation. Photosynthesis may be temporally impaired, but the cells respond by the production of secondary Cars, which can shield their chloroplasts against excessive irradiation or quench reactive oxygen species.

Identification of astaxanthin diglucoside diesters from snow alga Chlamydomonas nivalis by liquid chromatography–atmospheric pressure chemical ionization mass spectrometry

A method is described for the identification of astaxanthin glucoside esters from snow alga Chlamydomonas nivalis by means of liquid chromatography–mass spectrometry with atmospheric pressure chemical ionization (LC–MS/APCI). The method is based on the use of preparative HPLC and subsequent identification of astaxanthin diglucoside diesters by microbore LC–MS/APCI. The combination of these two techniques was used to identify more than 100 molecular species. The astaxanthin diglucoside diester, i.e. (all- E)-[di-(6- O-oleoyl-β- d-glucopyranosyloxy)]-astaxanthin, was also synthesized to unambiguously confirm its structure.

Seasonal and diel changes in photosynthetic activity of the snow alga Chlamydomonas nivalis (Chlorophyceae) from Svalbard determined by pulse amplitude modulation fluorometry

The seasonal and diel dynamics of the physiological state and photosynthetic activity of the snow alga Chlamydomonas nivalis were investigated in a snowfield in Svalbard. The snow surface represents an environment with very high irradiation intensities along with stable low temperatures close to freezing point. Photosynthetic activity was measured using pulse amplitude modulation fluorometry. Three types of cell (green biflagellate vegetative cells, orange spores clustered by means of mucilaginous sheaths, and purple spores with thick cell walls) were found, all of them photosynthetically active. The pH of snow ranged between 5.0 and 7.5, and the conductivity ranged between 5 and 75 microS cm(-1). The temperature of snow was stable (-0.1 to +0.1 degrees C), and the incident radiation values ranged from 11 to 1500 micromol photons m(-2) s(-1). The photosynthetic activity had seasonal and diel dynamics. The Fv/Fm values ranged between 0.4 and 0.7, and generally declined over the course of the season. A dynamic response of Fv/Fm to the irradiance was recorded. According to the saturating photon fluence values Ek, the algae may have obtained saturating light as deep as 3 cm in the snow when there were higher-light conditions, whereas they were undersaturated at prevalent low light even if on the surface.

UV-absorbing mycosporine-like compounds in planktonic and benthic organisms from a high-mountain lake

We investigated the occurrence, concentration and composition of mycosporines (mycosporine-like amino acids, MAAs) in planktonic organisms and epilithic cyanobacteria from a clear water high-mountain lake situated in the Central Alps, Austria. Two bi-substituted MAAs were identified by HPLC in extracts made of 1996 plankton samples with 90 % aqueous methanol, i.e. asterina-330 (&= = 330 nm) and shinorine (km = 334 run). Extracts with 20 % aqueous methanol for 2 h at 45 “C revealed the additional presence of another MAA tentatively identified as palythine (&.,, = 320nm) in the 1998 planktonic assemblage. In the upper 3 m of the water column the total concentration of MAAs decreased exponentially with depth, but the maxima for both absolute and chlorophyll-a specific concentrations were. observed close to the bottom at 8.5 m depth. This was explained by the accumulation of MAAs in the copepod’cyclops abyssorum tatricus that stays in deep water, during daytime. The copepodite III stage contained the 3 MAAs found in phytoplankton but also the mono-substituted compound, mycosporine-glycine (mycosporine-gly: & = 310 nm). The concentration of MAAs in C. abyssorum tatricus was highest for shinorine (1.45 % of the dry weight) and lowest for mycosporine-gly (0.02 % of the dry weight). Epilithic cyanobacteria had a more diverse MAA spectrum than plankton, and produced not only asterina-330 and shinorine but also palythinol (&, = 332 nm), mycosporine-gly and two unidentified compounds with &,,, = 330 and 340nm. The composition and also the relative abundance of the cyanobacterial MAAs changed with depth. Mycosporine-gly was found at the lakeshore where Gloeocapsa sp. dominates, but it was absent at 0.5 and 2.5 m depth dominated by Schizothrix sp. and Tolipothrix sp., respectively. We could not detect any MAAs in the cysts of the red snow alga Chlamydomonas nivalis, which develops on top of the winter cover shortly be’ Authors’ addresses: University of Innsbruck, InstitutGof Zoology and Limnology, Technikerstr. 25, A-6020 Innsbruck, Austria. E-mail: ruben.sommaruga@uibk.ac.at. 2 Max-Planck Institute for Marine Microbiology, Celsiusstr. 1, 28359 Bremen, Germany. * Author for correspondence. 0003-9136/99/0144-0255 $3.75

Phenolic compounds and antioxidant properties in the snow alga Chlamydomonas nivalis after exposure to UV light

The snow alga Chlamydomonas nivalis was collected from the Sierra Nevada, California, USA, and examined for its ability to produce phenolic compounds, free proline, and provide antioxidant protection factor in response to UV-A and UV-C light. Exposure of C. nivalis cells to UV-A light (365nm) for 5 days resulted in a 5–12% increase in total phenolics, where as exposure to UV-C light (254 nm) resulted in a 12–24% increase in phenolics after 7 days of exposure. Free proline was not affected by UV-A, but increased markedly after UV-C exposure. A three-fold increase in free proline occurred within two days after exposure to UV-C, but then dropped as cells became bleached. Antioxidant protection factor (PF) increased after treatment of cells with UV-A and remained constant throughout UV-C exposure. Spectral analysis of algal extracts revealed a decrease in absorption in the 215–225 nm region, short-term (2day) stimulation of pigment at 280 nm, and an increase in carotenoids (473 nm), after exposure to UV-A. Snow alga exposed to UV-C light had a different spectrum from that of UV-A exposed cells, i.e. an enhancement of three major peaks at 220, 260, and 280 nm, and loss of absorption in the carotenoid region.We report that UV light exposure, especially in the UV-C range, can stimulate phenolic-antioxidant production in aplanospores of C. nivalis effecting biochemical pathways related to proline metabolism.

RED SNOW CAUSED BY A NEW SPECIES OF TRACHELOMONAS(1) (2).

Trachelomonas kolii sp. nov., a member of the Euglenophyta, was the causative agent of red snow near Mt. Bachelor, Oregon. The cells are uniflagellate or without flagella and, in general, exhibit the characteristics of the genus Trachelomonas. The lorica, when present, is transparent, elongate, reticulate, and has a funnel-shaped collar at the anterior end. The protoplast is completely filled with red pigment. Red spherical resting stages that appear similar to the resting stages of the ubiquitous snow alga Chlamydomonas nivalis (Bauer) Wille are described.