Glycerolipid profile differences between perennial and annual stem zones in the perennial model plant Arabis alpina.

The perennial life style is a successful ecological strategy, and Arabis alpina is a recently developed model Brassicaceae species for studying it. One aspect, poorly investigated until today, concerns the differing patterns of allocation, storage, and metabolism of nutrients between perennials and annuals and the yet unknown signals that regulate this process. A. alpina has a complex lateral stem architecture with a proximal vegetative perennial (PZ) and a distal annual flowering zone (AZ) inside the same stems. Lipid bodies (LBs) with triacylglycerols (TAGs) accumulate in the PZ. To identify potential processes of lipid metabolism linked with the perennial lifestyle, we analyzed lipid species in the PZ versus AZ. Glycerolipid fractions, including neutral lipids with mainly TAGs, phospholipids, and glycolipids, were present at higher levels in the PZ as compared to AZ or roots. Concomitantly, contents of specific long-chain and very long-chain fatty acids increased during formation of the PZ. Corresponding gene expression data, gene ontology term enrichment, and correlation analysis with lipid species pinpoint glycerolipid-related genes to be active during the development of the PZ. Possibilities that lipid metabolism genes may be targets of regulatory mechanisms specifying PZ differentiation in A. alpina are discussed.

Cytokinin-promoted secondary growth and nutrient storage in the perennial stem zone of Arabis alpina.

Perennial plants maintain their lifespan through several growth seasons. Arabis alpina serves as a model Brassicaceae species to study perennial traits. Lateral stems of A. alpina have a proximal vegetative zone with a dormant bud zone and a distal senescing seed-producing inflorescence zone. We addressed how this zonation is distinguished at the anatomical level, whether it is related to nutrient storage and which signals affect the zonation. We found that the vegetative zone exhibits secondary growth, which we termed the perennial growth zone (PZ). High-molecular-weight carbon compounds accumulate there in cambium and cambium derivatives. Neither vernalization nor flowering were requirements for secondary growth and the sequestration of storage compounds. The inflorescence zone with only primary growth, termed the annual growth zone (AZ), or roots exhibited different storage characteristics. Following cytokinin application cambium activity was enhanced and secondary phloem parenchyma was formed in the PZ and also in the AZ. In transcriptome analysis, cytokinin-related genes represented enriched gene ontology terms and were expressed at a higher level in the PZ than in the AZ. Thus, A. alpina primarily uses the vegetative PZ for nutrient storage, coupled to cytokinin-promoted secondary growth. This finding lays a foundation for future studies addressing signals for perennial growth.

Overexpression of the chloroplastic 2-oxoglutarate/malate transporter disturbs carbon and nitrogen homeostasis in rice.

The chloroplastic 2-oxaloacetate (OAA)/malate transporter (OMT1 or DiT1) takes part in the malate valve that protects chloroplasts from excessive redox poise through export of malate and import of OAA. Together with the glutamate/malate transporter (DCT1 or DiT2), it connects carbon with nitrogen assimilation, by providing 2-oxoglutarate for the GS/GOGAT (glutamine synthetase/glutamate synthase) reaction and exporting glutamate to the cytoplasm. OMT1 further plays a prominent role in C4 photosynthesis: OAA resulting from phosphoenolpyruvate carboxylation is imported into the chloroplast, reduced to malate by plastidic NADP-malate dehydrogenase, and then exported for transport to bundle sheath cells. Both transport steps are catalyzed by OMT1, at the rate of net carbon assimilation. To engineer C4 photosynthesis into C3 crops, OMT1 must be expressed in high amounts on top of core C4 metabolic enzymes. We report here high-level expression of ZmOMT1 from maize in rice (Oryza sativa ssp. indica IR64). Increased activity of the transporter in transgenic rice was confirmed by reconstitution of transporter activity into proteoliposomes. Unexpectedly, overexpression of ZmOMT1 in rice negatively affected growth, CO2 assimilation rate, total free amino acid content, tricarboxylic acid cycle metabolites, as well as sucrose and starch contents. Accumulation of high amounts of aspartate and the impaired growth phenotype of OMT1 rice lines could be suppressed by simultaneous overexpression of ZmDiT2. Implications for engineering C4 rice are discussed.

Ammonia inhibits energy metabolism in astrocytes in a rapid and glutamate dehydrogenase 2-dependent manner.

Astrocyte dysfunction is a primary factor in hepatic encephalopathy (HE) impairing neuronal activity under hyperammonemia. In particular, the early events causing ammonia-induced toxicity to astrocytes are not well understood. Using established cellular HE models, we show that mitochondria rapidly undergo fragmentation in a reversible manner upon hyperammonemia. Further, in our analyses, within a timescale of minutes, mitochondrial respiration and glycolysis were hampered, which occurred in a pH-independent manner. Using metabolomics, an accumulation of glucose and numerous amino acids, including branched chain amino acids, was observed. Metabolomic tracking of 15N-labeled ammonia showed rapid incorporation of 15N into glutamate and glutamate-derived amino acids. Downregulating human GLUD2 [encoding mitochondrial glutamate dehydrogenase 2 (GDH2)], inhibiting GDH2 activity by SIRT4 overexpression, and supplementing cells with glutamate or glutamine alleviated ammonia-induced inhibition of mitochondrial respiration. Metabolomic tracking of 13C-glutamine showed that hyperammonemia can inhibit anaplerosis of tricarboxylic acid (TCA) cycle intermediates. Contrary to its classical anaplerotic role, we show that, under hyperammonemia, GDH2 catalyzes the removal of ammonia by reductive amination of α-ketoglutarate, which efficiently and rapidly inhibits the TCA cycle. Overall, we propose a critical GDH2-dependent mechanism in HE models that helps to remove ammonia, but also impairs energy metabolism in mitochondria rapidly.

Downregulation of a Mitochondrial NAD+ Transporter (NDT2) Alters Seed Production and Germination in Arabidopsis.

Despite the fundamental importance of nicotinamide adenine dinucleotide (NAD+) for metabolism, the physiological roles of NAD+ carriers in plants remain unclear. We previously characterized the Arabidopsis thaliana gene (At1g25380), named AtNDT2, encoding a protein located in the mitochondrial inner membrane, which imports NAD+ from the cytosol using ADP and AMP as counter-exchange substrates for NAD+. Here, we further investigated the physiological roles of NDT2, by isolating a T-DNA insertion line, generating an antisense line and characterizing these genotypes in detail. Reduced NDT2 expression affected reproductive phase by reducing total seed yield. In addition, reduced seed germination and retardation in seedling establishment were observed in the mutant lines. Moreover, remarkable changes in primary metabolism were observed in dry and germinated seeds and an increase in fatty acid levels was verified during seedling establishment. Furthermore, flowers and seedlings of NDT2 mutants displayed upregulation of de novo and salvage pathway genes encoding NAD+ biosynthesis enzymes, demonstrating the transcriptional control mediated by NDT2 activity over these genes. Taken together, our results suggest that NDT2 expression is fundamental for maintaining NAD+ balance amongst organelles that modulate metabolism, physiology and developmental processes of heterotrophic tissues.

Aroma and quality of breads baked from old and modern wheat varieties and their prediction from genomic and flour-based metabolite profiles.

Bread aroma is the principal characteristic perceived by the consumer yet it is mostly disregarded in the product chain. The main aim of this study was to evaluate the potential to include bread aroma as a new target criterion into the wheat product chain. The objectives of our study were to (i) quantify the influence of genetic versus environmental factors on the bread aroma and quality characteristics, (ii) evaluate whether bread baked from modern wheat varieties differ in terms of aroma from those baked from old varieties, and (iii) compare genomic and metabolomic approaches for their efficiency to predict bread aroma and quality characteristics in a wheat breeding program. Agronomic characters as well as bread aroma and quality traits were assessed for 18 old and 22 modern winter wheat varieties evaluated at up to three locations in Germany. Metabolite profiles of all 120 flour samples were collected using a 7200 GC-QTOF. Considerable differences in the adjusted entry means for all examined bread aroma and quality characters were observed. For aroma, which was rated on a scale from 1 to 9, the adjusted entry means varied for the 40 wheat varieties between 3 and 8. In contrast, the aroma of bread prepared from old and modern wheat varieties did not differ significantly (P < 0.05). Bread aroma was not significantly (P < 0.05) correlated with grain yield, which suggested that it is possible to select for the former character in wheat breeding programs without reducing the gain of selection for the latter. Finally, we have shown that bread aroma can be better predicted using a combination of metabolite and SNP genotyping profiles instead of the SNP genotyping profile only. In conclusion, we have illustrated possibilities to increase the quality of wheat for consumers in the product chain.

The sugar transporter SWEET10 acts downstream of FLOWERING LOCUS T during floral transition of Arabidopsis thaliana.

BACKGROUND: Floral transition initiates reproductive development of plants and occurs in response to environmental and endogenous signals. In Arabidopsis thaliana, this process is accelerated by several environmental cues, including exposure to long days. The photoperiod-dependent promotion of flowering involves the transcriptional induction of FLOWERING LOCUS T (FT) in the phloem of the leaf. FT encodes a mobile protein that is transported from the leaves to the shoot apical meristem, where it forms part of a regulatory complex that induces flowering. Whether FT also has biological functions in leaves of wild-type plants remains unclear. RESULTS: In order to address this issue, we first studied the leaf transcriptomic changes associated with FT overexpression in the companion cells of the phloem. We found that FT induces the transcription of SWEET10, which encodes a bidirectional sucrose transporter, specifically in the leaf veins. Moreover, SWEET10 is transcriptionally activated by long photoperiods, and this activation depends on FT and one of its earliest target genes SUPPRESSOR OF CONSTANS OVEREXPRESSION 1 (SOC1). The ectopic expression of SWEET10 causes early flowering and leads to higher levels of transcription of flowering-time related genes in the shoot apex. CONCLUSIONS: Collectively, our results suggest that the FT-signaling pathway activates the transcription of a sucrose uptake/efflux carrier during floral transition, indicating that it alters the metabolism of flowering plants as well as reprogramming the transcription of floral regulators in the shoot meristem.

Regional hypothermia improves gastric microcirculatory oxygenation during hemorrhage in dogs.

Mild systemic hypothermia increases gastric mucosal oxygenation (µHbO2) during hemorrhagic shock in dogs. In the context of critical blood loss hypothermia might be fatal due to adverse side effects. Selective regional hypothermia might overcome these limitations. The aim of our study was to analyze the effects of regional gastric and oral mucosal hypothermia on µHbO2 and perfusion (µflow). In a cross-over study six anesthetized dogs were subjected to local oral and gastric mucosal hypothermia (34°C), or maintenance of local normothermia during normovolemia and hemorrhage (-20% blood volume). Macro- and microcirculatory variables were recorded continuously. During normovolemia, local hypothermia increased gastric microcirculatory flow (µflow) without affecting oxygenation (µHbO2) or oral microcirculation. During mild hemorrhagic shock gastric µHbO2 decreased from 72±2% to 38±3% in the normothermic group. This was attenuated by local hypothermia, where µHbO2 was reduced from 74±3% to 52±4%. Local perfusion, oral microcirculation and macrocirculatory variables were not affected. Selective local hypothermia improves gastric µHbO2 during hemorrhagic shock without relevant side effects. In contrast to systemic hypothermia, regional mucosal hypothermia did not affect perfusion and oxygen supply during hemorrhage. Thus, the increased µHbO2 during local hypothermia rather indicates reduced mucosal oxygen demand.

Exogenous vasopressin dose-dependently modulates gastric microcirculatory oxygenation in dogs via V1A receptor.

BACKGROUND: Hypercapnia improves gastric microcirculatory oxygenation (µHbO2) and increases vasopressin plasma levels, whereas V1A receptor blockade abolishes the increase of µHbO2. The aim of this study was to evaluate the effect of exogenous vasopressin (AVP) in increasing doses on microcirculatory perfusion and oxygenation and systemic hemodynamic variables. Furthermore, we evaluated the role of the vasopressin V1A receptor in mediating the effects. METHODS: In repetitive experiments, six anesthetized dogs received a selective vasopressin V1A receptor inhibitor ([Pmp1, Tyr (Me)2]-Arg8-Vasopressin) or sodium chloride (control groups). Thereafter, a continuous infusion of AVP was started with dose escalation every 30 min (0.001 ng/kg/min-1 ng/kg/min). Microcirculatory variables of the oral and gastric mucosa were measured with reflectance spectrometry, laser Doppler flowmetry, and incident dark field imaging. Transpulmonary thermodilution was used to measure systemic hemodynamic variables. AVP plasma concentrations were measured during baseline conditions and 30 min after each dose escalation. RESULTS: During control conditions, gastric µHbO2 did not change during the course of experiments. Infusion of 0.001 ng/kg/min and 0.01 ng/kg/min AVP increased gastric µHbO2 to 87 ± 4% and 87 ± 6%, respectively, compared to baseline values (80 ± 7%), whereas application of 1 ng/kg/min AVP strongly reduced gastric µHbO2 (59 ± 16%). V1A receptor blockade prior to AVP treatment abolished these effects on µHbO2. AVP dose-dependently enhanced systemic vascular resistance (SVR) and decreased cardiac output (CO). After prior V1A receptor blockade, SVR was reduced and CO increased (0.1 ng/kg/min + 1 ng/kg/min AVP). CONCLUSIONS: Exogenous AVP dose-dependently modulates gastric µHbO2, with an increased µHbO2 with ultra-low dose AVP. The effects of AVP on µHbO2 are abolished by V1A receptor inhibition. These effects are independent of a modulation of systemic hemodynamic variables.

FlySilico: Flux balance modeling of Drosophila larval growth and resource allocation.

Organisms depend on a highly connected and regulated network of biochemical reactions fueling life sustaining and growth promoting functions. While details of this metabolic network are well established, knowledge of the superordinate regulatory design principles is limited. Here, we investigated by iterative wet lab and modeling experiments the resource allocation process during the larval development of Drosophila melanogaster. We chose this system, as survival of the animals depends on the successful allocation of their available resources to the conflicting processes of growth and storage metabolite deposition. First, we generated "FlySilico", a curated metabolic network of Drosophila, and performed time-resolved growth and metabolite measurements with larvae raised on a holidic diet. Subsequently, we performed flux balance analysis simulations and tested the predictive power of our model by simulating the impact of diet alterations on growth and metabolism. Our predictions correctly identified the essential amino acids as growth limiting factor, and metabolic flux differences in agreement with our experimental data. Thus, we present a framework to study important questions of resource allocation in a multicellular organism including process priorization and optimality principles.

The mitochondrial NAD+ transporter (NDT1) plays important roles in cellular NAD+ homeostasis in Arabidopsis thaliana.

Nicotinamide adenine dinucleotide (NAD+ ) is an essential coenzyme required for all living organisms. In eukaryotic cells, the final step of NAD+ biosynthesis is exclusively cytosolic. Hence, NAD+ must be imported into organelles to support their metabolic functions. Three NAD+ transporters belonging to the mitochondrial carrier family (MCF) have been biochemically characterized in plants. AtNDT1 (At2g47490), focus of the current study, AtNDT2 (At1g25380), targeted to the inner mitochondrial membrane, and AtPXN (At2g39970), located in the peroxisomal membrane. Although AtNDT1 was presumed to reside in the chloroplast membrane, subcellular localization experiments with green fluorescent protein (GFP) fusions revealed that AtNDT1 locates exclusively in the mitochondrial membrane in stably transformed Arabidopsis plants. To understand the biological function of AtNDT1 in Arabidopsis, three transgenic lines containing an antisense construct of AtNDT1 under the control of the 35S promoter alongside a T-DNA insertional line were evaluated. Plants with reduced AtNDT1 expression displayed lower pollen viability, silique length, and higher rate of seed abortion. Furthermore, these plants also exhibited an increased leaf number and leaf area concomitant with higher photosynthetic rates and higher levels of sucrose and starch. Therefore, lower expression of AtNDT1 was associated with enhanced vegetative growth but severe impairment of the reproductive stage. These results are discussed in the context of the mitochondrial localization of AtNDT1 and its important role in the cellular NAD+ homeostasis for both metabolic and developmental processes in plants.

Local gastric RAAS inhibition improves gastric microvascular perfusion in dogs.

During circulatory shock, gastrointestinal microcirculation is impaired, especially via activation of the renin-angiotensin-aldosterone system. Therefore, inhibition of the renin-angiotensin-aldosterone system might be beneficial in maintaining splanchnic microcirculation. The aim of this study was to analyze whether locally applied losartan influences gastric mucosal perfusion (µflow, µvelo) and oxygenation (µHbO2) without systemic hemodynamic changes. In repetitive experiments six anesthetized dogs received 30 mg losartan topically on the oral and gastric mucosa during normovolemia and hemorrhage (-20% blood volume). Microcirculatory variables were measured with reflectance spectrometry, laser Doppler flowmetry and incident dark field imaging. Transpulmonary thermodilution and pulse contour analysis were used to measure systemic hemodynamic variables. Gastric barrier function was assessed via differential absorption of inert sugars. During normovolemia, losartan increased gastric µflow from 99 ± 6 aU to 147 ± 17 aU and µvelo from 17 ± 1 aU to 19 ± 1 aU. During hemorrhage, losartan did not improve µflow. µvelo decreased from 17 ± 1 aU to 14 ± 1 aU in the control group. Application of losartan did not significantly alter µvelo (16 ± 1 aU) compared to the control group and to baseline levels (17 ± 1 aU). No effects of topical losartan on macrohemodynamic variables or microcirculatory oxygenation were detected. Gastric microcirculatory perfusion is at least partly regulated by local angiotensin receptors. Topical application of losartan improves local perfusion via vasodilation without significant effects on systemic hemodynamics. During mild hemorrhage losartan had minor effects on regional perfusion, probably because of a pronounced upstream vasoconstriction.

Corrigendum: Knock-Down of the Phosphoserine Phosphatase Gene Effects Rather N- Than S-Metabolism in Arabidopsis thaliana.

[This corrects the article DOI: 10.3389/fpls.2018.01830.].

Control of Drosophila Growth and Survival by the Lipid Droplet-Associated Protein CG9186/Sturkopf.

Lipid droplets (LDs) are the universal cellular storage organelles for esterified neutral lipids. The increasing number of characterized LD-associated proteins attained LDs with hitherto unexpected functions on top of their classical role as energy depot. Here, we characterize the LD-associated protein CG9186 of Drosophila by a CRISPR/Cas9-derived mutant fly line. While the mutant flies only showed a mild triacylglycerol storage phenotype, they were highly protected from desiccation stress, likely linked to a reduced locomotor activity and altered cuticular hydrocarbons. Both parameters depend on juvenile hormone (JH) signaling. Together with an observed interaction between CG9186 and JH-degrading enzymes, our results suggest that CG9186 participates in endocrine physiology regulation. In support of this hypothesis, CG9186 mutant flies show an altered expression of JH target genes and fail to adjust their developmental rate to dietary yeast-to-sugar ratio changes. Our results thus link LDs to organismic physiology regulation.

Nitrate or ammonium: Influences of nitrogen source on the physiology of a green alga.

In freshwaters, algal species are exposed to different inorganic nitrogen (Ni) sources whose incorporation varies in biochemical energy demand. We hypothesized that due to the lesser energy requirement of ammonium ( NH 4 + )-use, in contrast to nitrate ( NO 3 - )-use, more energy remains for other metabolic processes, especially under CO2- and phosphorus (Pi) limiting conditions. Therefore, we tested differences in cell characteristics of the green alga Chlamydomonas acidophila grown on NH 4 + or NO 3 - under covariation of CO2 and Pi-supply in order to determine limitations, in a full-factorial design. As expected, results revealed higher carbon fixation rates for NH 4 + -grown cells compared to growth with NO 3 - under low CO2 conditions. NO 3 - -grown cells accumulated more of the nine analyzed amino acids, especially under Pi-limited conditions, compared to cells provided with NH 4 + . This is probably due to a slower protein synthesis in cells provided with NO 3 - . In contrast to our expectations, compared to NH 4 + -grown cells NO 3 - -grown cells had higher photosynthetic efficiency under Pi-limitation. In conclusion, growth on the Ni-source NH 4 + did not result in a clearly enhanced Ci-assimilation, as it was highly dependent on Pi and CO2 conditions (replete or limited). Results are potentially connected to the fact that C. acidophila is able to use only CO2 as its inorganic carbon (Ci) source.

Multiomics resolution of molecular events during a day in the life of Chlamydomonas.

The unicellular green alga Chlamydomonas reinhardtii displays metabolic flexibility in response to a changing environment. We analyzed expression patterns of its three genomes in cells grown under light-dark cycles. Nearly 85% of transcribed genes show differential expression, with different sets of transcripts being up-regulated over the course of the day to coordinate cellular growth before undergoing cell division. Parallel measurements of select metabolites and pigments, physiological parameters, and a subset of proteins allow us to infer metabolic events and to evaluate the impact of the transcriptome on the proteome. Among the findings are the observations that Chlamydomonas exhibits lower respiratory activity at night compared with the day; multiple fermentation pathways, some oxygen-sensitive, are expressed at night in aerated cultures; we propose that the ferredoxin, FDX9, is potentially the electron donor to hydrogenases. The light stress-responsive genes PSBS, LHCSR1, and LHCSR3 show an acute response to lights-on at dawn under abrupt dark-to-light transitions, while LHCSR3 genes also exhibit a later, second burst in expression in the middle of the day dependent on light intensity. Each response to light (acute and sustained) can be selectively activated under specific conditions. Our expression dataset, complemented with coexpression networks and metabolite profiling, should constitute an excellent resource for the algal and plant communities.

NADP-MALIC ENZYME 1 Affects Germination after Seed Storage in Arabidopsis thaliana.

Aging decreases the quality of seeds and results in agricultural and economic losses. The damage that occurs at the biochemical level can alter the seed physiological status. Although loss of viability has been investigated frequently, little information exists on the molecular and biochemical factors involved in seed deterioration and loss of viability. Oxidative stress has been implicated as a major contributor to seed deterioration, and several pathways are involved in protection against this. In this study, we show that seeds of Arabidopsis thaliana lacking a functional NADP-MALIC ENZYME 1 (NADP-ME1) have reduced seed viability relative to the wild type. Seeds of the NADP-ME1 loss-of-function mutant display higher levels of protein carbonylation than those of the wild type. NADP-ME1 catalyzes the oxidative decarboxylation of malate to pyruvate with the simultaneous production of CO2 and NADPH. Upon seed imbibition, malate and amino acids accumulate in embryos of aged seeds of the NADP-ME1 loss-of-function mutant compared with those of the wild type. NADP-ME1 expression is increased in imbibed aged as compared with non-aged seeds. NADP-ME1 activity at testa rupture promotes normal germination of aged seeds. In seedlings of aged seeds, NADP-ME1 is specifically active in the root meristematic zone. We propose that NADP-ME1 activity is required for protecting seeds against oxidation during seed dry storage.

Efficient 2-phosphoglycolate degradation is required to maintain carbon assimilation and allocation in the C4 plant Flaveria bidentis.

Photorespiration is indispensable for oxygenic photosynthesis since it detoxifies and recycles 2-phosphoglycolate (2PG), which is the primary oxygenation product of Rubisco. However, C4 plant species typically display very low rates of photorespiration due to their efficient biochemical carbon-concentrating mechanism. Thus, the broader relevance of photorespiration in these organisms remains unclear. In this study, we assessed the importance of a functional photorespiratory pathway in the C4 plant Flaveria bidentis using knockdown of the first enzymatic step, namely 2PG phosphatase (PGLP). The isolated RNAi lines showed strongly reduced amounts of PGLP protein, but distinct signs of the photorespiratory phenotype only emerged below 5% residual PGLP protein. Lines with this characteristic were stunted in growth, had strongly increased 2PG content, exhibited accelerated leaf senescence, and accumulated high amounts of branched-chain and aromatic amino acids, which are both characteristics of incipient carbon starvation. Oxygen-dependent gas-exchange measurements consistently suggested the cumulative impairment of ribulose-1,5-bisphosphate regeneration with increased photorespiratory pressure. Our results indicate that photorespiration is essential for maintaining high rates of C4 photosynthesis by preventing the 2PG-mediated inhibition of carbon utilization efficiency. However, considerably higher 2PG accumulation can be tolerated compared to equivalent lines of C3 plants due to the differential distribution of specific enzymatic steps between the mesophyll and bundle sheath cells.

Cold Acclimation of the Thermoacidophilic Red Alga Galdieria sulphuraria: Changes in Gene Expression and Involvement of Horizontally Acquired Genes.

Galdieria sulphuraria is a unicellular red alga that lives in hot, acidic, toxic metal-rich, volcanic environments, where few other organisms survive. Its genome harbors up to 5% of genes that were most likely acquired through horizontal gene transfer. These genes probably contributed to G.sulphuraria's adaptation to its extreme habitats, resulting in today's polyextremophilic traits. Here, we applied RNA-sequencing to obtain insights into the acclimation of a thermophilic organism towards temperatures below its growth optimum and to study how horizontally acquired genes contribute to cold acclimation. A decrease in growth temperature from 42�C/46�C to 28�C resulted in an upregulation of ribosome biosynthesis, while excreted proteins, probably components of the cell wall, were downregulated. Photosynthesis was suppressed at cold temperatures, and transcript abundances indicated that C-metabolism switched from gluconeogenesis to glycogen degradation. Folate cycle and S-adenosylmethionine cycle (one-carbon metabolism) were transcriptionally upregulated, probably to drive the biosynthesis of betaine. All these cold-induced changes in gene expression were reversible upon return to optimal growth temperature. Numerous genes acquired by horizontal gene transfer displayed temperature-dependent expression changes, indicating that these genes contributed to adaptive evolution in G.sulphuraria.

The Photorespiratory BOU Gene Mutation Alters Sulfur Assimilation and Its Crosstalk With Carbon and Nitrogen Metabolism in Arabidopsis thaliana.

This study was aimed at elucidating the significance of photorespiratory serine (Ser) production for cysteine (Cys) biosynthesis. For this purpose, sulfur (S) metabolism and its crosstalk with nitrogen (N) and carbon (C) metabolism were analyzed in wildtype Arabidopsis and its photorespiratory bou-2 mutant with impaired glycine decarboxylase (GDC) activity. Foliar glycine and Ser contents were enhanced in the mutant at day and night. The high Ser levels in the mutant cannot be explained by transcript abundances of genes of the photorespiratory pathway or two alternative pathways of Ser biosynthesis. Despite enhanced foliar Ser, reduced GDC activity mediated a decline in sulfur flux into major sulfur pools in the mutant, as a result of deregulation of genes of sulfur reduction and assimilation. Still, foliar Cys and glutathione contents in the mutant were enhanced. The use of Cys for methionine and glucosinolates synthesis was reduced in the mutant. Reduced GDC activity in the mutant downregulated Calvin Cycle and nitrogen assimilation genes, upregulated key enzymes of glycolysis and the tricarboxylic acid (TCA) pathway and modified accumulation of sugars and TCA intermediates. Thus, photorespiratory Ser production can be replaced by other metabolic Ser sources, but this replacement deregulates the cross-talk between S, N, and C metabolism.