Carboxylase Activity Research Articles

ERCC6L2 Disease and Shwachman-Diamond Syndrome Exhibit Alterations in Metabolomes in Patient Fibroblasts

Introduction Metabolomics is an emerging tool to study metabolic alterations in cancers. It has also the potential to identify clinically relevant biomarkers for diagnosis, risk appraisal, and targeted drug development. In hematology, metabolic reprogramming is essential for normal regulation of hematopoiesis. Association of discrete metabolic alterations with explicit somatic mutations have also been reported in hematological malignancies. We have previously observed glutaminase (GLS) overexpression and improved oxygen consumption rate with excess glutamine in low glucose conditions in patient-derived fibroblasts in ERCC6L2 disease (unpublished). This prompted us to study metabolomic alterations in ERCC6L2 disease and Shwachman-Diamond syndrome (SDS) induced strictly by the germline ERCC6L2 and SBDS mutations. Both inherited bone marrow failure (BMF) syndromes carry considerable tendency of somatic TP53 mutagenesis predisposing patients to a high risk myeloid malignancy. Our aim here was to identify potential drug targets by targeted metabolomics. Methods We examined patient-derived skin fibroblasts (ERCC6L2 disease, n=4; SDS, n=4) and healthy controls (n=4) and performed targeted metabolomics with U-13C-D-glucose and U-13C-L-glutamine labeling. Fibroblasts were seeded on six-well plates and on the following day exposed to 13C-glucose (25mM) or 13C-glutamine (2mM) label containing medium for 30 or 120 minutes. Next, metabolites were extracted and quantified with liquid chromatography mass spectrometer. The results were normalized to total metabolite intensity per sample. Results In 13C-glucose labeled samples, we detected statistically significantly lower intensity of pyruvate M+3 in both ERCC6L2 and SDS fibroblasts compared to control at 30 minutes, which persisted at 120 minutes in SDS cells. In the TCA cycle, malate M+3 labeling indicates pyruvate carboxylase (PC) activity and M+2 labeling pyruvate dehydrogenase activity (PDH) (Figure 1). We found a significantly higher portion of malate M+3 than M+2 in SDS cells at all time points, suggesting lower PDH and higher PC activity in SDS cells. Additionally, we detected higher levels of alanine M+3 in SDS samples. As pyruvate is a source for alanine synthesis, we suspect an alteration in pyruvate metabolism; potentially to overcome challenges in pyruvate oxidation due to either decrease in the respiratory chain function or PDH activity. In 13C-glutamine labeled samples, we detected statistically significantly higher intensity of glutamine M+5 and M+4 in ERCC6L2 samples at 30 minutes compared to control and SDS cells. Glutamine is first converted to glutamate by glutaminase (GLS), after which it enters the TCA cycle as ketoglutarate (KG) (Figure 1). In addition to glutamine, we detected an increase, although not statistically significant, in glutamate M+5 and M+4, as well as KG M+5, in ERCC6L2 cells at 30 minutes. A similar trend was observed in the subsequent TCA cycle intermediates succinic acid M+4, fumarate M+4, and malate M+4. At 120 minutes, there was no difference in metabolite intensities between the groups, suggesting a potentially faster glutamine uptake by ERCC6L2 cells. Conclusions Our analysis using non-hematopoietic tissue provides understanding of metabolic weak links not confounded by somatic events occurring early in the disease course in ERCC6L2 and SDS patients. Our findings imply increased glutamine utilization in ERCC6L2 cells and altered pyruvate metabolism in SDS cells. Therefore, these metabolic pathways are attractive drug targets (summarized in Figure 1). Further studies are needed to determine their potential in supporting the compromised hematopoiesis to reduce the stress for somatic mutagenesis.

Effects of Vine Water Status on Malate Metabolism and γ-Aminobutyric Acid (GABA) Pathway-Related Amino Acids in Marselan (Vitis vinifera L.) Grape Berries.

Malic acid is the predominant organic acid in grape berries, and its content is affected by abiotic factors such as temperature (fruit zone microclimate) and water (vine water status). The objectives of this study were to explore the potential mechanisms behind the effects of vine water status on the biosynthesis and degradation of berry malic acid and the potential downstream effects on berry metabolism. This study was conducted over two growing seasons in 2021 and 2022, comprising three watering regimes: no water stress (CK), light water stress (LWS), and moderate water stress (MWS). Compared to CK, a significantly higher level of malic acid was found in berries from the MWS treatment when the berry was still hard and green (E-L 33) in both years. However, water stress reduced the malic acid content at the ripe berry harvest (E-L 38) stage. The activities of NAD-malate dehydrogenase (NAD-MDH) and pyruvate kinase (PK) were enhanced by water stress. Except for the E-L 33 stage, the activity of phosphoenolpyruvate carboxylase (PEPC) was reduced by water stress. The highest phosphoenolpyruvate carboxykinase (PEPCK) activity was observed at the berry veraison (E-L 35) stage and coincided with the onset of a decrease in the malate content. Meanwhile, the expression of VvPEPCK was consistent with its enzyme activity. This study showed that water stress changed the content of some free amino acids (GABA, proline, leucine, aspartate, and glutamate), two of which (glutamate and GABA) are primary metabolites of the GABA pathway.

Effects of polyethylene terephthalate microplastics on cell growth, intracellular products and oxidative stress of Scenedesmus sp.

Polyethylene terephthalate (PET) has been widely utilized in the synthesis of textile materials and packaging of foods and beverages. In recent years, it has been commonly detected in the form of microplastics (MPs) in wastewater. However, the effects of PET MPs on microalgal intracellular products and their interrelationships have been poorly investigated. In this study, the microalgae Scenedesmus sp. Strain H-1 was exposed to PET MPs to explore their effects on the growth, intracellular products (such as lipids, carbohydrates, and proteins), and antioxidative defense systems of Scenedesmus sp. The results demonstrated that PET MPs significantly reduced Scenedesmus sp. cell growth, with a maximum inhibition rate of 38.25% in the 500 mg L−1 treatment group. PET MPs had negative effects on glucose and nitrate utilization rates and reduced intracellular carbohydrates, intracellular proteins, and photosynthetic pigments. Surprisingly, PET MPs reduced acetyl-CoA carboxylase activity but induced lipid accumulation in microalgae. In addition, PET MPs significantly decreased the essential linoleic acid concentration and increased the palmitic acid content, resulting in reduced biodiesel quality. PET MPs induced the production of reactive oxygen species and malondialdehyde as well as the activities of superoxide dismutase and catalase. The results of the PCA indicated that the response mechanism of Scenedesmus sp. to PET MPs exposure was synergistic. This study provides fundamental data on the impact of MPs on the intracellular products of microalgae.

Waterlogging restricted cotton fiber elongation by reducing osmolyte accumulation and cell wall biosynthesis

Waterlogging severely threatens the formation of cotton fiber quality, especially the length. However, the underlying mechanism is still incompletely understood. Here, the experiment (control: soil relative water content at (75 ± 5)%; treatment: waterlogging for 6 days) was conducted during the flowering and boll forming stage using two upland cotton cultivars (Dexiamian 1 and Yuzaomian 9110) to investigate the effects of waterlogging on fiber development from both transcriptomic and physiological levels. The transcriptome analysis of 10 days post anthesis cotton fiber showed that waterlogging significantly regulated the expression of genes associated with fiber elongation events including osmotic solutes accumulation, fiber cell wall biosynthesis, and cell wall remodeling and loosening. Waterlogging decreased the sucrose input into cotton fiber, resulting in reduced sucrose and hexose contents. Under waterlogging, the accumulation of malate and K+ in both cultivars was reduced by decreasing phosphoenolpyruvate carboxylase (PEPC) activity and potassium transporter genes (GhPOT) expression, respectively. Thus, the total concentration of osmolytes was reduced, which dampened cotton fiber elongation. Due to reduced sucrose synthase activity and uridine diphosphate glucose content, the cellulose biosynthesis was limited, leading to decreased fiber cell wall accumulation, which was adverse to fiber elongation. Besides, genes responsible for cell wall remodeling and loosening, including xyloglucan endotransglucosylase/hydrolase genes (GhXTH), α-expansin genes (GhEXPA), and pectin methylesterase genes (GhPME) were also responsive to waterlogging, but in different ways between cultivars. Overall, the shortened cotton fiber could be attributed to reduced osmolytes accumulation and cell wall biosynthesis, whilst the effects of waterlogging on cotton fiber cell wall loosening require further exploration.

A bacteria-derived tetramerized protein ameliorates nonalcoholic steatohepatitis in mice via binding and relocating acetyl-coA carboxylase

Increased de novo lipogenesis (DNL) is a major feature of nonalcoholic steatohepatitis (NASH). None of the drugs targeting the catalytic activity of acetyl-CoA carboxylase (ACC), the rate-limiting enzyme in the DNL process, have been approved by the FDA. Whether cytosolic ACC1 can be regulated spatially remains to be explored. Herein, we find that streptavidin (SA), which is a bacterium-derived tetrameric protein, forms cytosolic condensates and efficiently induces a spatial re-localization of ACC1 in liver cells, concomitant with inhibited lipid accumulation. Both SA tetrameric structure and multivalent protein interaction are required for condensate formation. Interestingly, the condensates are further characterized as gel-like membraneless organelle (SAGMO) and significantly restrict the cytosolic dispersion of ACC1 and fatty acid synthase. Notably, AAV-mediated delivery of SA partially blocks mouse liver DNL and ameliorates NASH without eliciting hypertriglyceridemia. In summary, our study shows that insulating lipogenesis-related proteins by SAGMO might be effective for NASH treatment.

Gibberellic acid (GA) role in acetyl-coA carboxylase enzyme regulation and in improving oil palm yield

Seaweed specifically, Sargassum sp. is known to contains a boosting hormone growth that has been promoted plant growth and yield due to the containing of auxin, gibberelic acid (GA) and cytokinine, and also some amino acids especially glutamic acid. Those composition could be used as an booster of palm oil production which related to acetyl co-A carboxylase activity (ACC). ACC is the rate determination step in fatty acid accumulation, and becomes active through dephosphorylation of some serine residues that induced by magnesium and glutamate. Moreover, ACC was regulated by AtWRI1 and AtWRI1-TCP4 interaction, a mechanism that allow fine-tuning of the oil biosynthetic pathway. In this research we conducted gene expression experiments, and molecular docking analyses to study the possible mechanism of seaweed composition stimulating oil accumulation in the oil palm. Further analysis was conducted to ensure whether the interaction between TCP4 and candidate inhibitors were able to phosphorylate TCP4 and decrease its activity. GA application resulted in the increase of oil accumulation in 1 month after application, although in the second month the oil accumulation showed decreasing. Increase of oil accumulation in the first month in line with the increase of the expression of ACC in 3rd and 5th weeks. Meanwhile, TCP4 showed decrease expression that resulted in the increase of the WRI1 in 5th week. From this result, it was indicated that GA application could block the TCP4, so it could not interact with WRI1, resulted in the expression of WRI1 and ACC. This interaction stimulates the oil accumulation in oil palm.

Gibberellic acid (GA) role in acetyl-coA carboxylase enzyme regulation and in improving oil palm yield

Seaweed specifically, Sargassum sp. is known to contains a boosting hormone growth that has been promoted plant growth and yield due to the containing of auxin, gibberelic acid (GA) and cytokinine, and also some amino acids especially glutamic acid. Those composition could be used as an booster of palm oil production which related to acetyl co-A carboxylase activity (ACC). ACC is the rate determination step in fatty acid accumulation, and becomes active through dephosphorylation of some serine residues that induced by magnesium and glutamate. Moreover, ACC was regulated by AtWRI1 and AtWRI1-TCP4 interaction, a mechanism that allow fine-tuning of the oil biosynthetic pathway. In this research we conducted gene expression experiments, and molecular docking analyses to study the possible mechanism of seaweed composition stimulating oil accumulation in the oil palm. Further analysis was conducted to ensure whether the interaction between TCP4 and candidate inhibitors were able to phosphorylate TCP4 and decrease its activity. GA application resulted in the increase of oil accumulation in 1 month after application, although in the second month the oil accumulation showed decreasing. Increase of oil accumulation in the first month in line with the increase of the expression of ACC in 3rd and 5th weeks. Meanwhile, TCP4 showed decrease expression that resulted in the increase of the WRI1 in 5th week. From this result, it was indicated that GA application could block the TCP4, so it could not interact with WRI1, resulted in the expression of WRI1 and ACC. This interaction stimulates the oil accumulation in oil palm.

Lycopene Alleviates the Adverse Effects of Feeding High-Lipid Diets to Hybrid Grouper (♀Epinephelus fuscoguttatus ×♂E. lanceolatus).

It has been found that high-lipid diets (HLDs) disrupt lipid metabolism in fish, leading to an excessive accumulation of lipids in various tissues of the fish body. The objective of this study was to investigate if the inclusion of lycopene (LCP) in an HLD may mitigate the adverse consequences of excessive dietary lipid intake in hybrid grouper (♀ Epinephelus fuscoguttatus × ♂ E. lanceolatus). The experimental design incorporated a control group (L0), which was administered a diet consisting of 42% protein and 16% lipid. The diets for groups L1, L2, and L3 were developed by augmenting the control diet with 100, 200, and 400 mg/kg LCP, respectively. The duration of the trial spanned a period of 42 days. The results of the study showed that the weight gain rate (WGR) and protein efficiency ratio (PER) of the three LCP treatment groups (L1, L2, and L3) tended to increase and then decrease, with a significant increase in WGR and PER in L2 (P < 0.05). Visceral somatic index and hepatic somatic index tended to decrease and then increase in all treatment groups, with a significant decrease in L2 (P < 0.05). In serum dietary LCP significantly reduced triglyceride (TG), total cholesterol (TC), and low-density lipoprotein (LDL) content and significantly increased high-density lipoprotein (HDL) content (P < 0.05). In the liver, dietary LCP reduced TC, TG, and very LDL levels and improved lipoprotein lipase, hepatic lipase, fatty acid (FA) synthetase, and acetyl-CoA carboxylase activities. The number and area of hepatic lipid droplets decreased significantly with increasing LCP content. In the liver, the addition of appropriate levels of LCP significantly upregulated lipoprotein lipase (lpl) and peroxisome proliferator-activated receptor α (pparα). In summary, dietary LCP improves growth and reduces lipid deposition in the liver of hybrid grouper by increasing lipolytic metabolism and decreasing FA synthesis. Under the experimental conditions, the fitted curve analysis showed that the recommended LCP additions to the high lipid diet for juvenile hybrid grouper were 200-300 mg/kg.

Different Dietary Sources of Selenium Alleviate Hepatic Lipid Metabolism Disorder of Heat-Stressed Broilers by Relieving Endoplasmic Reticulum Stress.

As global warming continues, the phenomenon of heat stress (HS) in broilers occurs frequently. The alleviating effect of different selenium (Se) sources on HS-induced hepatic lipid metabolism disorders in broilers remains unclear. This study compared the protective effects of four Se sources (sodium selenite; selenium yeast; selenomethionine; nano-Se) on HS-induced hepatic lipid metabolism disorder and the corresponding response of selenotranscriptome in the liver of broilers. The results showed that HS-induced liver injury and hepatic lipid metabolism disorder, which were reflected in the increased activity of serum alanine aminotransferase (ALT), the increased concentration of triacylglycerol (TG) and total cholesterol (TC), the increased activity of acetyl-CoA carboxylase (ACC), diacylglycerol O-acyltransferase (DGAT) and fatty acid synthase (FAS), and the decreased activity of hepatic lipase (HL) in the liver. The hepatic lipid metabolism disorder was accompanied by the increased mRNA expression of lipid synthesis related-genes, the decreased expression of lipidolysis-related genes, and the increased expression of endoplasmic reticulum (ER) stress biomarkers (PERK, IRE1, ATF6, GRP78). The dietary supplementation of four Se sources exhibited similar protective effects. Four Se sources increased liver Se concentration and promoted the expression of selenotranscriptome and several key selenoproteins, enhanced liver antioxidant capacity and alleviated HS-induced ER stress, and thus resisted the hepatic lipid metabolism disorders of broilers exposed to HS. In conclusion, dietary supplementation of four Se sources (0.3 mg/kg) exhibited similar protective effects on HS-induced hepatic lipid metabolism disorders of broilers, and the protective effect is connected to the relieving of ER stress.

Overexpressing plant ferredoxin-like protein enhances photosynthetic efficiency and carbohydrates accumulation in Phalaenopsis.

Crassulacean acid metabolism (CAM) is one of three major models of carbon dioxide assimilation pathway with better water-use efficiency and slower photosynthetic efficiency in photosynthesis. Previous studies indicated that the gene of sweet pepper plant ferredoxin-like protein (PFLP) shows high homology to the ferredoxin-1(Fd-1) family that belongs to photosynthetic type Fd and involves in photosystem I. It is speculated that overexpressing pflp in the transgenic plant may enhance photosynthetic efficiency through the electron transport chain (ETC). To reveal the function of PFLP in photosynthetic efficiency, pflp transgenic Phalaenopsis, a CAM plant, was generated to analyze photosynthetic markers. Transgenic plants exhibited 1.2-folds of electron transport rate than that of wild type (WT), and higher CO2 assimilation rates up to 1.6 and 1.5-folds samples at 4pm and 10pm respectively. Enzyme activity of phosphoenolpyruvate carboxylase (PEPC) was increased to 5.9-folds in Phase III, and NAD+-linked malic enzyme (NAD+-ME) activity increased 1.4-folds in Phase IV in transgenic plants. The photosynthesis products were analyzed between transgenic plants and WT. Soluble sugars contents such as glucose, fructose, and sucrose were found to significantly increase to 1.2, 1.8, and 1.3-folds higher in transgenic plants. The starch grains were also accumulated up to 1.4-folds in transgenic plants than that of WT. These results indicated that overexpressing pflp in transgenic plants increases carbohydrates accumulation by enhancing electron transport flow during photosynthesis. This is the first evidence for the PFLP function in CAM plants. Taken altogether, we suggest that pflp is an applicable gene for agriculture application that enhances electron transport chain efficiency during photosynthesis.

Effects of exogenous melatonin on sugar and organic acid metabolism in early-ripening peach fruits.

To evaluated the effects melatonin (MT) on the sugar and acid metabolism of early-ripening peach fruits, the concentration of 150 μmol/L MT was sprayed on the leaves of peach trees. MT increased the contents of total soluble sugar and sucrose in peach fruits during the whole ripening period, and increased the contents of glucose and sorbitol at the mature stage. During the whole ripening period, MT also increased the activities of sucrose synthase, sucrose phosphate synthase, neutral invertase, and acidic invertase and the relative expression levels of sucrose synthase, sucrose phosphate synthase, neutral invertase, and acidic invertase genes, while decreased the activity of sorbitol oxidase and the relative expression level of sorbitol dehydrogenase to some extent. Moreover, MT decreased the contents of total organic acid, malic acid, and citric acid at mature stage. At mature stage, MT decreased the activities of citrate synthetase and phosphoenolpyruvate carboxylase and the relative expression levels of citrate synthetase and phosphoenolpyruvate carboxylase genes, while increased the relative expression levels of Nicotinamide adenine dinucleotide phosphate (NADP+)-malic enzyme, malate dehydrogenase, and aconitase genes. Therefore, MT promotes the sugar accumulation and organic acid degradation in early-ripening peach fruits.

Comparative metabolomics analysis investigating the impact of melatonin-enriched diet on energy metabolism in the crayfish, Cherax destructor.

Melatonin is a multifunctional bioactive molecule present in almost all organisms and has been gradually used in the aquaculture industry in recent years. Energy metabolism is an essential process for individuals to maintain their life activities; however, the process through which melatonin regulates energy metabolism in aquatic animals remains unclear. The present study aimed to conduct a comprehensive analysis of the regulatory mechanism of melatonin for energy metabolism in Cherax destructor by combining metabolomics analysis with the detection of the key substance content, enzymatic activity, and gene expression levels in the energy metabolism process after culturing with dietary melatonin supplementation for 8weeks. Our results showed that dietary melatonin increased the content of glycogen, triglycerides, and free fatty acids; decreased lactate levels; and promoted the enzymatic activity of pyruvate kinase (PK), malate dehydrogenase (MDH), and acetyl-CoA carboxylase. The results of gene expression analysis showed that dietary melatonin also increased the expression levels of hexokinase, PK, MDH, lactate dehydrogenase, lipase, and fatty acid synthase genes. The results of metabolomics analysis showed that differentially expressed metabolites were significantly enriched in lysine degradation and glycerophospholipid metabolism. In conclusion, our study demonstrates that dietary melatonin increased oxidative phosphorylation, improved glucose utilization, and promoted storage of glycogen and lipids in C. destructor. These lipids are used not only for energy storage but also to maintain the structure and function of cell membranes. Our results further add to the understanding of the mechanisms of energy regulation by melatonin in crustaceans.

Effects of dietary melatonin on growth performance, nutrient composition, and lipid metabolism of Pacific white shrimp (Penaeus vannamei)

The goal of this study was to investigate the effects of dietary melatonin (MT) on the growth physiology and lipid metabolism of Penaeus vannamei (P. vannamei). We prepared six experimental diets with varying MT levels (0 (control), 22.5, 41.2, 82.7, 165.1, 329.2 mg/kg) and fed six groups of shrimp with these diets for 2 months. The highest survival, body length growth rate, weight gain, and specific growth rates occurred in shrimp that were fed the diet containing 82.7 mg/kg MT, and the highest levels of docosahexaenoic acid (DHA) and arachidonic acid (ARA) were also found in this treatment group. Compared with the control group, there were no significant differences in moisture, crude protein, and ash contents in shrimp fed with MT, but the crude lipid content was significantly decreased in the high concentration MT supplemented group. Compared to the control group, shrimp fed 82.7 mg/kg MT had better morphology and structure of the hepatopancreas, whereas the hepatopancreas tissues were damaged in shrimp from the high dose MT groups. Analysis of lipid metabolism-related enzymes and genes in the hepatopancreas showed that dietary MT greatly decreased fatty acid synthase and acetyl-CoA carboxylase activities but increased carnitine palmitoyltransferase 1 activity. At the transcriptional level, dietary MT reduced hepatopancreas fat accumulation by upregulating lipid oxidation genes and downregulating lipid synthesis genes. A broken-line model revealed that the ideal MT concentration was in the range of 59.59–71.97 mg/kg. We found that MT supplementation improved P. vannamei growth performance, enhanced fatty acid β oxidation, and prevented fat deposition in the hepatopancreas.

Metabolic heterogeneity of tissue-resident macrophages in homeostasis and during helminth infection

Tissue-resident macrophage populations constitute a mosaic of phenotypes, yet how their metabolic states link to the range of phenotypes and functions in vivo is still poorly defined. Here, using high-dimensional spectral flow cytometry, we observe distinct metabolic profiles between different organs and functionally link acetyl CoA carboxylase activity to efferocytotic capacity. Additionally, differences in metabolism are evident within populations from a specific site, corresponding to relative stages of macrophage maturity. Immune perturbation with intestinal helminth infection increases alternative activation and metabolic rewiring of monocyte-derived macrophage populations, while resident TIM4+ intestinal macrophages remain immunologically and metabolically hyporesponsive. Similar metabolic signatures in alternatively-activated macrophages are seen from different tissues using additional helminth models, but to different magnitudes, indicating further tissue-specific contributions to metabolic states. Thus, our high-dimensional, flow-based metabolic analyses indicates complex metabolic heterogeneity and dynamics of tissue-resident macrophage populations at homeostasis and during helminth infection.

Effect of carbon and nitrogen concentrations on lipid accumulation and regulation of acetyl-CoA carboxylase in Papiliotrema laurentii.

Biodiesel is an interesting alternative to petroleum diesel as it is renewable, biodegradable, and has a low pollutant content. Yeast oils can be used for biodiesel production instead of edible oils, mitigating the use of arable land and water for biodiesel production. Maximum lipid accumulation is reached at 48h of cultivation by the oleaginous yeast Papiliotrema laurentii UFV-1. Nevertheless, the effects of carbon and nitrogen concentrations on lipid accumulation, as well as the regulation of lipid metabolism in this yeast are still not well-characterised. Therefore, this work evaluated the effects of carbon and nitrogen concentrations on the lipid accumulation in P. laurentti, the expression of the ACC gene, and the activity of the enzyme acetyl-CoA carboxylase (ACCase) in different carbon:nitrogen ratios (C:N) and glucose concentrations. The variation of ammonium sulfate concentration did not affect the growth and lipid accumulation in P. laurentii UFV-1. On the other hand, glucose concentration remarkably influenced biomass and lipid production by this yeast. Therefore, the carbon concentration is more important than the nitrogen concentration for lipid production by P. laurentii UFV-1. Importantly, the levels of both ACC gene expression and ACCase activity were maximum during the late-exponential growth phase and decreased after reaching the highest lipid contents, which was easier evidenced during the accumulation and maximum lipid levels. As such, the reduction of ACCase enzyme activity seems to be related to the decrease in the expression level of the ACC gene.

The isolation and functional identification of a phosphoenolpyruvate carboxykinase gene from Saccharina japonica

Similar to other macroalgae, Saccharina japonica has CO2-concentrating mechanism (CCM) that allows high photosynthesis efficiency and elevates biomass. Phosphoenolpyruvate carboxykinase (PEPCK) in cytoplasm is an essential component of CCM. However, no reports on cytosolic PEPCK of S. japonica are available. In this study, the full-length cDNA sequence of a PEPCK gene (SjPCK1) predicted to be localized in cytoplasm, was screened from the full-length transcriptome of S. japonica gametophytes and identified by RT-PCR. The complete cDNA sequence of SjPCK1 was 2174 bp in length, which encompassed an open reading frame (ORF) of 1734 bp, a 5′- untranslated region (UTR) of 216 bp and a 3′-UTR of 224 bp. In parallel, the genomic DNA of SjPCK1 was 21 294 bp in length, characterized by the presence of 11 introns and 12 exons. It encoded a protein of 577 amino acids with a molecular weight of 63 kD and an isoelectric point of 8.47. Amino acid sequence alignment showed that the functional sites of PEPCK were highly conserved in the selected species. Phylogenetic analysis and sequence characterization showed that SjPCK1 was an ATP-dependent PEPCK. SjPCK1 gene was expressed by using Escherichia coli prokaryotic expression system, and the SjPCK1 protein with His 6 tag (rSjPCK1) was 81.93 kD in molecular weight. Enzyme activity assay results showed that the specific activity of carboxylase and decarboxylase of rSjPCK1 was 1.91 ± 0.29 U/mg prog and 11.3 ± 1.97 U/mg prog, respectively. These findings provide valuable molecular and biochemical insights for a further analysis of the role of cytosolic PEPCK in the storage of inorganic carbon in S. japonica.

Phosphorus and naphthalene acetic acid increased the seed yield by regulating carbon and nitrogen assimilation of flax.

To evaluate the impact of phosphorus (P) combined with exogenous NAA on flax yield, enhance flax P utilization efficiency and productivity, minimize resource inputs and mitigate negative environmental and human effects. Therefore, it is crucial to comprehend the physiological and biochemical responses of flax to P and naphthylacetic acid (NAA) in order to guide future agronomic management strategies for increasing seed yield. A randomized complete block design trial was conducted under semi-arid conditions in Northwest China, using a factorial split-plot to investigate the effects of three P (0, 67.5, and 135.0kg P2O5 ha-1) and three exogenous spray NAA levels (0, 20, and 40 mg NAA L-1) on sucrose phosphate synthase (SPS) and diphosphoribulose carboxylase (Rubisco) activities as well as nitrogen (N) and P accumulation and translocation in flax. Results indicated that the SPS and Rubisco activities, N and P accumulation at flowering and maturity along with assimilation and translocation post-flowering, fruiting branches per plant, tillers per plant, capsules per plant, and seed yield were 95, 105, 14, 27, 55, 15, 13, 110, 103, 82, 16, 61, 8, and 13% greater in the P treatments compared to those in the zero P treatment, respectively. Moreover, those characteristics were observed to be greater with exogenous spray NAA treatments compared to that no spray NAA treatment. Additionally, the maximum SPS and Rubisco activities, N and P accumulation, assimilation post-flowering and translocation, capsules per plant, and seed yield were achieved with the application of 67.5kg P2O5 ha-1 with 20 mg NAA L-1. Therefore, these findings demonstrate that the appropriate combination of P fertilizer and spray NAA is an effective agronomic management strategy for regulating carbon and nitrogen assimilation by maintaining photosynthetic efficiency in plants to increase flax productivity.

The potential of seaweed extracts as a biostimulant for improving salt stress tolerance of Solanum lycopersicum L

The present investigation aimed to examine the effects of seaweed extracts (SWE) on tomato plants under saline conditions (50 mM of NaCl). The current findings have shown that the SWE obtained from Jania rubens (J. rubens) gave better results compared to those prepared from the other two seaweeds; Dictyota dichotoma (D. dichotoma) and Gelidium sesquipedale (G. sesquipedale). Indeed, under salt stress, the application of fresh seaweed at 2% significantly (p ≤ 0.05) increased the plant height (26.28 cm ± 0.95) compared with the non-treated stressed control (17.65 cm ± 1.04). Based on these results the effect of J. rubens on some biological and biochemical parameters of tomato plants was evaluated. In fact, increased contents of chlorophylls, total soluble sugars (TSS), and total free amino acids (TFAA) were observed in seaweed-treated plants compared to the control plants. Moreover, the treatment of stressed plants with SWE revealed a significant (p ≤ 0.05) improvement in antioxidant enzyme activities such as superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione S-transferase (GST). The extract also showed an increase in the enzyme activities of glutamate dehydrogenase (GDH) and phosphoenolpyruvate carboxylase (PEPC), which are implicated in the nitrogen and carbon metabolism, respectively. Furthermore, the analysis of the chemical composition of the liquid seaweed extracts (fresh and dried) has shown that they contain bioactive compounds such as polyphenols, flavonoids, soluble sugars, and amino acids, which can be responsible for the tomato's tolerance to salinity. Therefore, the SWE tested in the present study can be used as a biostimulant to enhance the salt stress tolerance of tomato plants.

Dietary choline activates the Ampk/Srebp signaling pathway and decreases lipid levels in Pacific white shrimp (Litopenaeus vannamei)

An 8-week feeding trial was conducted in Pacific white shrimp (Litopenaeus vannamei) to evaluate the effects of dietary choline supplementation on choline transport and metabolism, hepatopancreas histological structure and fatty acid profile, and regulation of lipid metabolism. Six isonitrogenous and isolipidic diets were formulated to contain different choline levels of 2.91 (basal diet), 3.85, 4.67, 6.55, 10.70 and 18.90 g/kg, respectively. A total of 960 shrimp (initial weight, 1.38 ± 0.01 g) were distributed randomly into twenty-four 250-L cylindrical fiber-glass tanks, with each diet assigned randomly to 4 replicate tanks. The results indicated that dietary choline significantly promoted the deposition of choline, betaine and carnitine (P < 0.05). The diameters and areas of R cells, total lipid and triglyceride contents in hepatopancreas, and triglyceride and non-esterified fatty acid contents in hemolymph were negatively correlated with dietary choline level. The contents of functional fatty acids in hepatopancreas, the activity of acetyl-CoA carboxylase (Acc), and the mRNA expression of fas, srebp and acc were highest in shrimp fed the diet containing 4.67 g/kg choline, and significantly higher than those fed the diet containing 2.91 g/kg, the lowest level of choline (P < 0.05). The number of R cells, content of very low-density lipoprotein (VLDL), activities of carnitine palmitoyl-transferase (Cpt1), lipoprotein lipase and hepatic lipase, and the mRNA expression levels of cpt1, fabp, fatp, ldlr, and ampk in hepatopancreas increased significantly as dietary choline increased (P < 0.05). In addition, hepatopancreas mRNA expression levels of ctl1, ctl2, oct1, badh, bhmt, ck, cept, and cct were generally up-regulated as dietary choline level increased (P < 0.01). In conclusion, dietary choline promoted the deposition of choline and its metabolites by up-regulating genes related to choline transport and metabolism. Moreover, appropriate dietary choline level promoted the development of hepatopancreas R cells and maintained the normal accumulation of lipids required for development, while high dietary choline not only promoted hepatopancreas lipid export by enhancing VLDL synthesis, but also promoted fatty acid β-oxidation and inhibited de novo fatty acid synthesis by activating the Ampk/Srebp signaling pathway. These findings provided further insight and understanding of the mechanisms by which dietary choline regulated lipid metabolism in L. vannamei.

Pyruvate anaplerosis is a targetable vulnerability in persistent leukaemic stem cells

Deregulated oxidative metabolism is a hallmark of leukaemia. While tyrosine kinase inhibitors (TKIs) such as imatinib have increased survival of chronic myeloid leukaemia (CML) patients, they fail to eradicate disease-initiating leukemic stem cells (LSCs). Whether TKI-treated CML LSCs remain metabolically deregulated is unknown. Using clinically and physiologically relevant assays, we generate multi-omics datasets that offer unique insight into metabolic adaptation and nutrient fate in patient-derived CML LSCs. We demonstrate that LSCs have increased pyruvate anaplerosis, mediated by increased mitochondrial pyruvate carrier 1/2 (MPC1/2) levels and pyruvate carboxylase (PC) activity, in comparison to normal counterparts. While imatinib reverses BCR::ABL1-mediated LSC metabolic reprogramming, stable isotope-assisted metabolomics reveals that deregulated pyruvate anaplerosis is not affected by imatinib. Encouragingly, genetic ablation of pyruvate anaplerosis sensitises CML cells to imatinib. Finally, we demonstrate that MSDC-0160, a clinical orally-available MPC1/2 inhibitor, inhibits pyruvate anaplerosis and targets imatinib-resistant CML LSCs in robust pre-clinical CML models. Collectively these results highlight pyruvate anaplerosis as a persistent and therapeutically targetable vulnerability in imatinib-treated CML patient-derived samples.