Adenosylhomocysteinase Research Articles

Molecular Mechanism of NL13 Peptide of Adenosyl Homocysteinase Against ER Stress through Nrf2 Signaling Cascade

Molecular Mechanism of NL13 Peptide of Adenosyl Homocysteinase Against ER Stress through Nrf2 Signaling Cascade

Effect of icariin on depressive behaviour in rat pups. Evidences for its mechanism of action by integrating network pharmacology, metabolomics and gut microbiota composition

BackgroundPrenatal stress (PS) can cause cognitive disorder and a range of psychological illnesses, including anxiety and depression. Icariin (ICA) has shown promising effects in improving PS-induced depressive behaviour. However, its mechanism of action remains unclear. PurposeThis study was performed to reveal the key targets, metabolites and gut microbiota for ICA in improving depressive behaviour in PS rat pups. MethodsA prenatal restraint stress animal model was established for Sprague–Dawley (SD) rats in late pregnancy. Male pups were randomly divided into six groups: no stress group (NS), PS group, PS + saline group (PS_S), PS + high-dose ICA group (ICAH, 80 mg/kg*day), PS + low-dose ICA group (ICAL, 40 mg/kg*day) and PS + fluoxetine group (FLU, 10 mg/kg*day). The depressive behaviour of each group of rat pups was evaluated using open field test, forced swimming test and sucrose preference test. Different metabolites were identified using untargeted metabolomics of serum and faeces, and metabolic pathways were analyzed through MetaboAnalyst. Targets for ICA acting on depression were determined after network pharmacology was applied. An integrated network of network pharmacology and metabolomics were constructed using Cytoscape software, and molecular docking were performed to verify the interactions between ICA and key targets. Finally, gut microbiota of rat pups in each group were analyzed after 16S rDNA sequencing. ResultsPS could cause rat pups to exhibit depressive behaviour, and ICA could significantly improve this depressive behaviour. A total of 49 differential metabolites were found in serum and 23 differential metabolites were found in faeces, and 24 metabolites in serum and 6 metabolites in faeces could be reversed following ICA administration. Integrated analysis focused on five key targets (i.e. adenosyl homocysteinase; medium-chain specific acyl-CoA dehydrogenase, mitochondrial; thymidine phosphorylase; cGMP-specific 3′,5′-cyclic phosphodiesterase and xanthine dehydrogenase/oxidase) and three metabolites (i.e. palmitoylcarnitine, methionine and hypoxanthine). Molecular docking indicated that ICA combined well with key targets. Gut microbiota analysis showed that g_Bacteroides, f_Bacteroidaceae and s_Lactobacillus reuteri were required for ICA to improve depressive behaviour. ConclusionIn this study, the antidepressant mechanism of ICA was clarified with a strategy of integrating metabolomics, network pharmacology and gut microbiota. ICA has a good effect on improving metabolism and increasing the abundance of probiotics in the intestine. The present research provided new insights into the anti-depressant mechanism of ICA.

Zika virus remodelled ER membranes contain proviral factors involved in redox and methylation pathways

Zika virus (ZIKV) has emerged as a global health issue, yet neither antiviral therapy nor a vaccine are available. ZIKV is an enveloped RNA virus, replicating in the cytoplasm in close association with ER membranes. Here, we isolate ER membranes from ZIKV-infected cells and determine their proteome. Forty-six host cell factors are enriched in ZIKV remodeled membranes, several of these having a role in redox and methylation pathways. Four proteins are characterized in detail: thioredoxin reductase 1 (TXNRD1) contributing to folding of disulfide bond containing proteins and modulating ZIKV secretion; aldo-keto reductase family 1 member C3 (AKR1C3), regulating capsid protein abundance and thus, ZIKV assembly; biliverdin reductase B (BLVRB) involved in ZIKV induced lipid peroxidation and increasing stability of viral transmembrane proteins; adenosylhomocysteinase (AHCY) indirectly promoting m6A methylation of ZIKV RNA by decreasing the level of S- adenosyl homocysteine and thus, immune evasion. These results highlight the involvement of redox and methylation enzymes in the ZIKV life cycle and their accumulation at virally remodeled ER membranes.

Adenosylhomocysteinase plays multiple roles in maintaining the identity and pluripotency of mouse embryonic stem cells†.

Adenosylhomocysteinase (AHCY), a key enzyme in the methionine cycle, is essential for the development of embryos and the maintenance of mouse embryonic stem cells (mESCs). However, the precise underlying mechanism of Ahcy in regulating pluripotency remains unclear. As the only enzyme that can hydrolyze S-adenosylhomocysteine in mammals, AHCY plays a critical role in the metabolic homeostasis, epigenetic remodeling, and transcriptional regulation. Here, we identified Ahcy as a direct target of OCT4 and unveiled that AHCY regulates the self-renewal and differentiation potency of mESCs through multiple mechanisms. Our study demonstrated that AHCY is required for the metabolic homeostasis of mESCs. We revealed the dual role of Ahcy in both transcriptional activation and inhibition, which is accomplished via the maintenance of H3K4me3 and H3K27me3, respectively. We found that Ahcy is required for H3K4me3-dependent transcriptional activation in mESCs. We also demonstrated that AHCY interacts with polycomb repressive complex 2 (PRC2), thereby maintaining the pluripotency of mESCs by sustaining the H3K27me3-regulated transcriptional repression of related genes. These results reveal a previously unrecognized OCT4-AHCY-PRC2 axis in the regulation of mESCs' pluripotency and provide insights into the interplay between transcriptional factors, cellular metabolism, chromatin dynamics and pluripotency regulation.

Multi-Omics Analysis Reveals Intricate Gene Networks Involved in Female Development in Melon.

Sexual differentiation is an important developmental phenomenon in cucurbits that directly affects fruit yield. The natural existence of multiple flower types in melon offers an inclusive structure for studying the molecular basis of sexual differentiation. The current study aimed to identify and characterize the molecular network involved in sex determination and female development in melon. Male and female pools separated by the F2 segregated generation were used for sequencing. The comparative multi-omics data revealed 551 DAPs and 594 DEGs involved in multiple pathways of melon growth and development, and based on functional annotation and enrichment analysis, we summarized four biological process modules, including ethylene biosynthesis, flower organ development, plant hormone signaling, and ubiquitinated protein metabolism, that are related to female development. Furthermore, the detailed analysis of the female developmental regulatory pathway model of ethylene biosynthesis, signal transduction, and target gene regulation identified some important candidates that might have a crucial role in female development. Two CMTs ((cytosine-5)-methyltransferase), one AdoHS (adenosylhomocysteinase), four ACSs (1-aminocyclopropane-1-carboxylic acid synthase), three ACOs (ACC oxidase), two ARFs (auxin response factor), four ARPs (auxin-responsive protein), and six ERFs (Ethylene responsive factor) were identified based on various female developmental regulatory models. Our data offer new and valuable insights into female development and hold the potential to offer a deeper comprehension of sex differentiation mechanisms in melon.

Metabolic profiling stratifies colorectal cancer and reveals adenosylhomocysteinase as a therapeutic target

The genomic landscape of colorectal cancer (CRC) is shaped by inactivating mutations in tumour suppressors such as APC, and oncogenic mutations such as mutant KRAS. Here we used genetically engineered mouse models, and multimodal mass spectrometry-based metabolomics to study the impact of common genetic drivers of CRC on the metabolic landscape of the intestine. We show that untargeted metabolic profiling can be applied to stratify intestinal tissues according to underlying genetic alterations, and use mass spectrometry imaging to identify tumour, stromal and normal adjacent tissues. By identifying ions that drive variation between normal and transformed tissues, we found dysregulation of the methionine cycle to be a hallmark of APC-deficient CRC. Loss of Apc in the mouse intestine was found to be sufficient to drive expression of one of its enzymes, adenosylhomocysteinase (AHCY), which was also found to be transcriptionally upregulated in human CRC. Targeting of AHCY function impaired growth of APC-deficient organoids in vitro, and prevented the characteristic hyperproliferative/crypt progenitor phenotype driven by acute deletion of Apc in vivo, even in the context of mutant Kras. Finally, pharmacological inhibition of AHCY reduced intestinal tumour burden in ApcMin/+ mice indicating its potential as a metabolic drug target in CRC.

Abstract 1747: Targeting metabolic and epigenetic programs to re-sensitize glioblastoma to chemotherapy

Abstract Treatment options for glioblastoma (GBM) are limited. Prognosis remains dismal, with an 18 month on average survival rate following diagnosis due to treatment resistance and disease recurrence. The goal of this project is to investigate hallmarks of cancer progression that contribute to temozolomide (TMZ) resistance, a first tine treatment for GBM. Two signaling pathways were investigated in TMZ-sensitive and -resistant GBM cell lines and in primary and recurrent patient-derived xenograft (PDX) tumor cells by genetically and pharmacologically inhibiting methionine adenosyltransferase 2A (MAT2A) and adenosylhomocysteinase (AHCY). Cell growth and survival were assessed by measuring protein expression of proliferation, oxidative stress and cell cycle arrest markers. EPIC array analysis and targeted bisulfite sequencing were conducted to identify changes in genome-wide and specific CpG island methylation. The Seahorse XF Analyzer measured mitochondrial respiratory capacity and oxidative metabolism. Induced pluripotent stem cell organoids were co-cultured with PDX tumor cells to determine if treatments mitigate tumor cell invasiveness. Compared to parental cells (PC), MAT2A gene expression was increased by 1.7-fold in acquired resistant and de novo resistant GBM cells (RC) [(transcript per million): PC, 7386 ± 412; RC, 12941 ± 1023; n=2; p=2.10e-8].Compared to TMZ-sensitive cells (TS), TMZ-resistant cells (TR) demonstrated a 56% increase in baseline oxygen consumption rate [(pmol/min): TS, 179 ± 6.7; TR, 279 ± 13; n=18; p=.0012] and 64% increase in maximal respiratory capacity [(pmol/min): TS, 403 ± 29; TR, 659 ± 35; n=6; p<.0001]. Both primary and recurrent GBM cells were subject to select MAT2A and AHCY inhibitor compounds and EC50s were determined. Recurrent cells demonstrated a vulnerability to TMZ-generated cell death both by genetically knocking down and pharmacologically inhibiting MAT2A and AHCY. MAT2A and AHCY contribute to TMZ resistance and recurrence by dysregulating methylation programs and upregulating antioxidant programs, respectively. These findings provide a foundation for developing novel combinatory therapeutic strategies and inform clinical studies intended to augment the practically negligible rate of remission for GBM and improve treatment outcomes for cancer patients. Citation Format: Emma Rowland, Jordan Walter, Anna Jermakowicz, Robert Suter, Rebecca Riggins, Nagi Ayad. Targeting metabolic and epigenetic programs to re-sensitize glioblastoma to chemotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1747.

337 Targeting metabolic and epigenetic programs to re-sensitize glioblastoma to chemotherapy

OBJECTIVES/GOALS: Treatment options for glioblastoma (GBM) are limited. Prognosis remains dismal, with an 18 month on average survival rate following diagnosis due to treatment resistance and disease recurrence. The goal of this project is to investigate hallmarks of cancer progression that contribute to temozolomide (TMZ) resistance, a first tine treatment for GBM. METHODS/STUDY POPULATION: Two signaling pathways were investigated in TMZ-sensitive and -resistant GBM cell lines and in primary and recurrent patient-derived xenograft (PDX) tumor cells by genetically and pharmacologically inhibiting methionine adenosyltransferase 2A (MAT2A) and adenosylhomocysteinase (AHCY). Cell growth and survival were assessed by measuring protein expression of proliferation, oxidative stress and cell cycle arrest markers. EPIC array analysis and targeted bisulfite sequencing were conducted to identify changes in genome-wide and specific CpG island methylation. The Seahorse XF Analyzer measured mitochondrial respiratory capacity and oxidative metabolism. Induced pluripotent stem cell organoids were co-cultured with PDX tumor cells to determine if treatments mitigate tumor cell invasiveness. RESULTS/ANTICIPATED RESULTS: Compared to parental cells (PC), MAT2A gene expression was increased by 1.7-fold in acquired resistant and de novo resistant GBM cells (RC) [(transcript per million): PC, 7386 ± 0.012; RC, 12925 ± 0.023; n=2; p=2.10e-8]. Compared to TMZ-sensitive cells (TS), TMZ-resistant cells (TR) demonstrated a 56% increase in baseline oxygen consumption rate [(pmol/min): TS, 179 ± 6.7; TR, 279 ± 13; n=18; p=.0012] and 64% increase in maximal respiratory capacity [(pmol/min): TS, 403 ± 29; TR, 659 ± 35; n=6; p DISCUSSION/SIGNIFICANCE: MAT2A and AHCY contribute to TMZ resistance and recurrence by dysregulating methylation programs and upregulating antioxidant programs, respectively. These findings provide a foundation for developing novel combinatory therapeutic strategies and inform clinical studies intended to increase remission and reduce recurrence for GBM patients.

Discovery of AHCY as an Off-Target of Doxorubicin by Integrative Analysis of Photoaffinity Labeling Chemoproteomics and Untargeted Metabolomics.

Target identification is critically important for understanding the mechanism of action of drugs. Here, we reported a new strategy for deconvolution of drug targets (or off-targets) with photoaffinity labeling chemoproteomics in combination with untargeted metabolomics by using doxorubicin (DOX) as a model. The DOX-derived photoaffinity probes were prepared and applied to capture DOX-interacting proteins in living cells. The captured DOX-interacting proteins were then identified by label-free quantitative proteomics. Totally, 151 significant proteins were identified with high confidence (fold change >4, p-value < 0.005). The gene ontology enrichment analysis suggested that the proteins were mainly involved in carbon metabolism, citrate cycle, fatty acid metabolism, and metabolic pathways. Therefore, untargeted metabolomics was applied to quantify the significantly altered metabolites in cells upon drug treatment. The pathway enrichment analysis suggested that DOX mainly interrupted with the processes of pyrimidine and purine metabolism, carbon metabolism, methionine metabolism, and phosphatidylcholine biosynthesis. Integrative analysis of chemoproteomics and metabolomics indicated that adenosylhomocysteinase (AHCY) is a new target (off-target) of DOX leading to the accumulation of S-adenosyl homocysteine. This deduced DOX target was confirmed by the cellular thermal shift assay, affinity competitive pull-down assay, biochemical assay, and siRNA knock down experiments. Our result suggested that AHCY is the uncovered off-target of DOX.

The long non-coding RNA lncMYOZ2 mediates an AHCY/MYOZ2 axis to promote adipogenic differentiation in porcine preadipocytes

Long non-coding RNAs (lncRNAs) play a vital role in regulating adipogenesis. However, the associated regulatory mechanisms have yet to be described in detail in pig. In this study, we demonstrate a critical role for lncMYOZ2 in adipogenesis from porcine preadipocytes. Specifically, lncMYOZ2 was more abundant in the adipose tissue of Mashen (fat-type) pigs than for Large White (lean-type) pigs, and knockdown of this lncRNA significantly inhibited the differentiation of porcine preadipocytes into adipocytes. Mechanistically, we used RNA pull-down and RIP assays to establish that lncMYOZ2 interacts with adenosylhomocysteinase (AHCY). Moreover, lncMYOZ2 knockdown increased promoter methylation of the target gene MYOZ2 and lowered its expression. Finally, we describe a positive regulatory role for MYOZ2 in adipogenesis. Collectively, these findings establish lncMYOZ2 as an important epigenetic regulator of adipogenesis via the aforementioned AHCY/MYOZ2 pathway, and provide insights into the role of lncRNAs in porcine adipose development.

Attenuation of SARS‐CoV‐2 replication and associated inflammation by concomitant targeting of viral and host cap 2'‐O‐ribose methyltransferases

The SARS‐CoV‐2 infection cycle is a multistage process that relies on functional interactions between the host and the pathogen. Here, we repurposed antiviral drugs against both viral and host enzymes to pharmaceutically block methylation of the viral RNA 2'‐O‐ribose cap needed for viral immune escape. We find that the host cap 2'‐O‐ribose methyltransferase MTr1 can compensate for loss of viral NSP16 methyltransferase in facilitating virus replication. Concomitant inhibition of MTr1 and NSP16 efficiently suppresses SARS‐CoV‐2 replication. Using in silico target‐based drug screening, we identify a bispecific MTr1/NSP16 inhibitor with anti‐SARS‐CoV‐2 activity in vitro and in vivo but with unfavorable side effects. We further show antiviral activity of inhibitors that target independent stages of the host SAM cycle providing the methyltransferase co‐substrate. In particular, the adenosylhomocysteinase (AHCY) inhibitor DZNep is antiviral in in vitro, in ex vivo, and in a mouse infection model and synergizes with existing COVID‐19 treatments. Moreover, DZNep exhibits a strong immunomodulatory effect curbing infection‐induced hyperinflammation and reduces lung fibrosis markers ex vivo. Thus, multispecific and metabolic MTase inhibitors constitute yet unexplored treatment options against COVID‐19.

Prognostic Value of Serum Exosomal AHCY Expression in Hepatitis B-Induced Liver Cirrhosis.

Objective: We aimed to investigate serum exosomal adenosylhomocysteinase (AHCY) expression in hepatitis B-induced liver cirrhosis (HBV-LC) patients and to determine the prognostic value of serum exosomal AHCY.Methods: We collected serum samples from 100 patients with chronic hepatitis B (CHB) and from 114 HBV-LC patients to test serum exosomal AHCY expression using ELISA.Results: Compared with the CHB and Grade A and B HBV-LC groups, the level of exosomal AHCY expression was significantly higher in the HBV-LC group [376.62 (291.50–448.02) vs. 248.12 (189.28–324.63), P > 0.001] and the Grade C HBV-LC group [408.70 (365.63–465.76) vs. 279.76 (215.16–336.07), P > 0.001], respectively. Serum exosomal AHCY expression and MELD score had a significant positive correlation (r = 0.844, P < 0.001). Survival curve analysis showed that patients with low exosomal AHCY expression had significantly longer survival than patients with high exosomal AHCY expression (P = 0.0038). The receiver operating characteristics (ROC) curve showed that the area under the curve (AUC) value for the mortality prediction ability of serum exosomal AHCY in HBV-LC patients was 0.921, which was higher than the values for the MELD score (AUC 0.815) and Child-Pugh classification (AUC 0.832), with a sensitivity and specificity of 93.41 and 76.00%, respectively.Conclusions: The serum exosomal AHCY level is a novel potential prognostic biomarker in HBV-LC patients, which may be great significance for the prognosis of HBV-LC patients.

AHCYL1 senses SAH to inhibit autophagy through interaction with PIK3C3 in an MTORC1-independent manner

ABSTRACT S-adenosyl-l-homocysteine (SAH), an amino acid derivative, is a key intermediate metabolite in methionine metabolism, which is normally considered as a harmful by-product and hydrolyzed quickly once formed. AHCY (adenosylhomocysteinase) converts SAH into homocysteine and adenosine. There are two other members in the AHCY family, AHCYL1 (adenosylhomocysteinase like 1) and AHCYL2 (adenosylhomocysteinase like 2). Here we define AHCYL1 function as a SAH sensor to inhibit macroautophagy/autophagy through PIK3C3. The C terminus of AHCYL1 interacts with SAH specifically and the interaction with SAH promotes the binding of the N terminus to the catalytic domain of PIK3C3, resulting in inhibition of PIK3C3. More importantly, this observation was further validated in vivo, indicating that SAH functions as a signaling molecule. Our study uncovers a new axis of SAH-AHCYL1-PIK3C3, which senses the intracellular level of SAH to inhibit autophagy in an MTORC1-independent manner. Abbreviations: ADOX: adenosine dialdehyde; AHCY: adenosylhomocysteinase; AHCYL1: adenosylhomocysteinase like 1; cLEU: cycloleucine; PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3; PtdIns3P: phosphatidylinositol-3-phosphate; SAH: S-adenosyl-l-homocysteine; SAM: S-adenosyl-l-methionine.

Coordinated expression of Jumonji and AHCY under OCT transcription factor control to regulate gene methylation in wood frogs during anoxia.

Wood frogs (Rana sylvatica) can survive extended periods of whole body freezing. Freezing imparts multiple stresses on cells that include anoxia and dehydration, but these can also be experienced as independent stresses. Under anoxia stress, energy metabolism is suppressed, and pro-survival pathways are prioritized to differentially regulate some transcription factors including OCT1 and OCT4. Jumonji C domain proteins (JMJD1A and JMJD2C) are hypoxia responsive demethylases whose expression is accelerated by OCT1 and OCT4 which act to demethylate genes related to the methionine cycle. The responses by these factors to 24h anoxia exposure and 4h aerobic recovery was analyzed in liver and skeletal muscle of wood frogs to assess their involvement in metabolic adaptation to oxygen limitation. Immunoblot results showed a decrease in JMJD1A levels under anoxia in liver and muscle, but an increase was observed in JMJD2C demethylase protein in anoxic skeletal muscle. Protein levels of adenosylhomocysteinase (AHCY) and methionine adenosyl transferase (MAT), enzymes of the methionine cycle, also showed an increase in the reoxygenated liver, whereas the levels decreased in muscle. A transcription factor ELISA showed a decrease in DNA binding by OCT1 in the reoxygenated liver and anoxic skeletal muscle, and transcript levels also showed tissue specific gene expression. The present study provides the first analysis of the role of the OCT1 transcription factor, associated proteins, and lysine demethylases in mediating responses to anoxia by wood frog tissues.

Functional and Pathological Roles of AHCY.

Adenosylhomocysteinase (AHCY) is a unique enzyme and one of the most conserved proteins in living organisms. AHCY catalyzes the reversible break of S-adenosylhomocysteine (SAH), the by-product and a potent inhibitor of methyltransferases activity. In mammals, AHCY is the only enzyme capable of performing this reaction. Controlled subcellular localization of AHCY is believed to facilitate local transmethylation reactions, by removing excess of SAH. Accordingly, AHCY is recruited to chromatin during replication and active transcription, correlating with increasing demands for DNA, RNA, and histone methylation. AHCY deletion is embryonic lethal in many organisms (from plants to mammals). In humans, AHCY deficiency is associated with an incurable rare recessive disorder in methionine metabolism. In this review, we focus on the AHCY protein from an evolutionary, biochemical, and functional point of view, and we discuss the most recent, relevant, and controversial contributions to the study of this enzyme.

S-adenosyl-l-homocysteine hydrolase links methionine metabolism to the circadian clock and chromatin remodeling.

Circadian gene expression driven by transcription activators CLOCK and BMAL1 is intimately associated with dynamic chromatin remodeling. However, how cellular metabolism directs circadian chromatin remodeling is virtually unexplored. We report that the S-adenosylhomocysteine (SAH) hydrolyzing enzyme adenosylhomocysteinase (AHCY) cyclically associates to CLOCK-BMAL1 at chromatin sites and promotes circadian transcriptional activity. SAH is a potent feedback inhibitor of S-adenosylmethionine (SAM)-dependent methyltransferases, and timely hydrolysis of SAH by AHCY is critical to sustain methylation reactions. We show that AHCY is essential for cyclic H3K4 trimethylation, genome-wide recruitment of BMAL1 to chromatin, and subsequent circadian transcription. Depletion or targeted pharmacological inhibition of AHCY in mammalian cells markedly decreases the amplitude of circadian gene expression. In mice, pharmacological inhibition of AHCY in the hypothalamus alters circadian locomotor activity and rhythmic transcription within the suprachiasmatic nucleus. These results reveal a previously unappreciated connection between cellular metabolism, chromatin dynamics, and circadian regulation.

Proteomic investigation of Zn-challenged rice roots reveals adverse effects and root physiological adaptation

AimsUnderstanding the molecular responses of plant roots to the challenging environment contributes to engineering plants with improved stress tolerance. However, little has been done to understand rice (Oryza sativa L.) roots response to the toxic concentration of zinc (Zn) at the proteomic level. This study explored proteomic responses of young rice roots 5–6 days after sowing to 765 μM of Zn in a hydroponic set-up after 4-day exposure.MethodsDye staining method and spectrophotometry were chosen for physiological response investigations. ICP-MS was used to determine metal content in rice seedlings. Two-dimensional gel electrophoresis was used for the proteomic study of root tissue.ResultsElevated Zn reduced Mg translocation to the shoots and Mn uptake, decreased plant growth, and increased cell death of roots. Zn stimulated the biosynthesis of glutathione but decreased ROS and malondialdehyde levels. In total, 42 Zn-responsive proteins were identified. Proteins involved in redox regulation, defense response, sulfur metabolism, and proteolysis were induced, while those of energy production and cell wall biogenesis were decreased.ConclusionRoots of O. sativa seedlings could tolerate certain Zn excess (765 μM Zn for 4 days) by stimulating nutrient recycling and glycolysis, and production of defensive metabolites and proteins to maintain ion and redox homeostasis. Besides, adenosylhomocysteinase (AHCY) down-regulation may contribute to balanced transmethylations, and methionine salvage pathway appears important for the adaptation to Zn. Our results provide some new insight into a complex metabolic response of rice roots to Zn stress, which is related to both stress and defense mechanisms.

O-GlcNAcylation regulates the methionine cycle to promote pluripotency of stem cells

Methionine metabolism is critical for the maintenance of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) pluripotency. However, little is known about the regulation of the methionine cycle to sustain ESC pluripotency. Here, we show that adenosylhomocysteinase (AHCY), an important enzyme in the methionine cycle, is critical for the maintenance and differentiation of mouse embryonic stem cells (mESCs). We show that mESCs exhibit high levels of methionine metabolism, whereas decreasing methionine metabolism via depletion of AHCY promotes mESCs to differentiate into the three germ layers. AHCY is posttranslationally modified with an O-linked β-N-acetylglucosamine sugar (O-GlcNAcylation), which is rapidly removed upon differentiation. O-GlcNAcylation of threonine 136 on AHCY increases its activity and is important for the maintenance of trimethylation of histone H3 lysine 4 (H3K4me3) to sustain mESC pluripotency. Blocking glycosylation of AHCY decreases the ratio of S-adenosylmethionine versus S-adenosylhomocysteine (SAM/SAH), reduces the level of H3K4me3, and poises mESC for differentiation. In addition, blocking glycosylation of AHCY reduces somatic cell reprogramming. Thus, our findings reveal a critical role of AHCY and a mechanistic understanding of O-glycosylation in regulating ESC pluripotency and differentiation.

Antioxidant molecular mechanism of adenosyl homocysteinase from cyanobacteria and its wound healing process in fibroblast cells.

An antioxidant molecule namely, adenosyl homocysteinase (AHc) was identified from the earlier constructed transcriptome database of Spirulina, where it was cultured in a sulphur deprived condition. From the AHc protein, a small peptide NL13 was identified using bioinformatics tools and was predicted to have antioxidant property. Further, the peptide was synthesised and its antioxidant mechanism was addressed at molecular level. NL13 was subjected to various antioxidant assays including DPPH assay, HARS assay, SARS Assay, NO assay and ABTS assay, where NL13 exhibited significant (P < 0.05) potential antioxidant activity compared to its antioxidant control, Trolox. Cytotoxicity was performed on Human whole blood and the cell viability was performed on VERO fibroblast cells. In both assays, it was found that NL13 did not exhibit any cytotoxic effect towards the cells. Further, the intracellular ROS was performed on Multimode reader followed by imaging on fluorescence microscope which showed scavenging activity even at lower concentration of NL13 (31.2 µM). An effective wound healing property of NL13 on VERO cells was confirmed by analysing the cell migration rate at two different time intervals (24 and 48 h). Overall, the study shows that NL13 peptide scavenges the intracellular oxidative stress.Electronic supplementary materialThe online version of this article (10.1007/s11033-020-05276-y) contains supplementary material, which is available to authorized users.

A murine model of wheat versus potato allergy: Patatin and 53kDa protein are the potential allergen from potato

Wheat allergy is the most common around the world as gluten is the potential allergen. People diagnosed with wheat allergy were mainly substitute with other novel food such as potato though it is also being reported for allergenic manifestations. Thus there is an increasing demand for developing a BALB/c mice model to empathize the allergic properties of potato protein and its fractions. Purified potato protein showed lower IgE-binding capacity (474.39 ± 0.6 ng/mL) even in higher concentration (30 mg/mL) compared to wheat gluten (1418.28 ± 0.17 ng/mL, 5mg/mL). Immediate active cutaneous hypersensitivity reaction, vascular leakage, intestinal permeability and lung’s inflammatory cell infiltration was also ascertained comparatively lower in potato protein than wheat gluten. Furthermore, patatin (43 kDa) and protease inhibitors (∼21 kDa) were purified and separated, and patatin exhibited higher hypersensitivity score than that of protease inhibitors. Immuno-detection assays indicated that patatin and 53 kDa protein in potato protein showed specific Ig-E binding capacity, and 53 kDa was adenosyl homocysteinase identified by LC–MS/MS.