Transferase Gene Research Articles

Association of histamine-N-methyl transferase gene polymorphisms with carnosine content in red-brown Korean native chickens.

Carnosine and anserine affect the meat flavor. The contents of carnosine and anserine in meat are affected by genetic and environmental factors. This study aimed to discover the single-nucleotide polymorphisms (SNPs) in the histamine-N-methyl transferase (HNMT) and histamine-N-methyl transferase-like (HNMT-like) genes and to associate them with the content of carnosine and anserine in Korean native chickenred brown line (KNC-R). This study used a total of 384 birds (males, n = 192; females, n = 192) aged 10 weeks old, for genotyping HNMT and HNMT-like genes. One synonymous SNP (rs29009298C/T) of the HNMT gene was genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods whereas four missense SNPs (rs734406537G/A; rs736514667A/G; rs15881680G/A and rs316765035T/C) of the HNMT gene, and one missense SNP rs737657949A/C of the HNMT-like gene were genotyped by PCR allele competitive extension (PACE) genotyping technology. Two-way analysis of variance of the R program was used to associate HNMT genotypes with the contents of carnosine and anserine in KNC-R chickens. There were significant associations (p<0.05) between the genotypes of the synonymous SNP:rs29009298C/T, missense SNP rs736514667A/G of the HNMT gene and the content of carnosine in KNC-Rs. This study also reported the sex effect on the carnosine content, where females had more content of carnosine compared to that of male KNC-R. Two SNPs (synonymous: rs735769522C/T) and missense: rs736514667A/G) in the HNMT gene might be used as genetic markers in the selection and breeding of chickens with better taste and high-flavored meat.

An in-silico analysis of OGT gene association with diabetes mellitus

O-GlcNAcylation is a nutrient-sensing post-translational modification process. This cycling process involves two primary proteins: the O-linked N-acetylglucosamine transferase (OGT) catalysing the addition, and the glycoside hydrolase OGA (O-GlcNAcase) catalysing the removal of the O-GlCNAc moiety on nucleocytoplasmic proteins. This process is necessary for various critical cellular functions. The O-linked N-acetylglucosamine transferase (OGT) gene produces the OGT protein. Several studies have shown the overexpression of this protein to have biological implications in metabolic diseases like cancer and diabetes mellitus (DM). This study retrieved 159 SNPs with clinical significance from the SNPs database. We probed the functional effects, stability profile, and evolutionary conservation of these to determine their fit for this research. We then identified 7 SNPs (G103R, N196K, Y228H, R250C, G341V, L367F, and C845S) with predicted deleterious effects across the four tools used (PhD-SNPs, SNPs&Go, PROVEAN, and PolyPhen2). Proceeding with this, we used ROBETTA, a homology modelling tool, to model the proteins with these point mutations and carried out a structural bioinformatics method– molecular docking– using the Glide model of the Schrodinger Maestro suite. We used a previously reported inhibitor of OGT, OSMI-1, as the ligand for these mutated protein models. As a result, very good binding affinities and interactions were observed between this ligand and the active site residues within 4Å of OGT. We conclude that these mutation points may be used for further downstream analysis as drug targets for treating diabetes mellitus.

Biochemical characterization and gene structure analysisof the 24-kDa glutathione transferase sigma from Taenia solium.

Taenia solium can cause human taeniasis and/or cysticercosis. The latter can in some instances cause human neurocysticercosis which is considered a priority in disease-control strategies and the prevention of mental health problems. Glutathione transferases are crucial for the establishment and long-term survival of T. solium; therefore, we structurally analyzed the 24-kDa glutathione transferase gene (Ts24gst) of T. solium and biochemically characterized its product. The gene promoter showed potential binding sites for transcription factors and xenobiotic regulatory elements. The gene consists of a transcription start site, four exons split by three introns, and a polyadenylation site. The gene architecture is conserved in cestodes. Recombinant Ts24GST (rTs24GST) was active and dimeric. Anti-rTs24GST serum showed slight cross-reactivity with human sigma-class GST. A 3D model of Ts24GST enabled identification of putative residues involved in interactions of the G-site with GSH and of the H-site with CDNB and prostaglandin D2. Furthermore, rTs24GST showed optimal activity at 45 °C and pH 9, as well as high structural stability in a wide range of temperatures and pHs. These results contribute to the better understanding of this parasite and the efforts directed to fight taeniasis/cysticercosis.

Uncovering the Role of Hydroxycinnamoyl Transferase in Boosting Chlorogenic Acid Accumulation in Carthamus tinctorius Cells under Methyl Jasmonate Elicitation.

Chlorogenic acids (CGAs) are bioactive compounds widely used in the food, pharmaceutical, and cosmetic industries. Carthamus tinctorius is an important economic crop, and its suspension cells are rich in CGAs. However, little is known about the biosynthesis and regulation of CGAs in Carthamus tinctorius cells. This study first elucidated the regulatory mechanism of CGA biosynthesis in methyl jasmonate (MeJA)-treated Carthamus tinctorius cells and the role of the MeJA-responsive hydroxycinnamoyl transferase (HCT) gene in enhancing their CGA accumulation. Firstly, temporal changes in intracellular metabolites showed that MeJA increased the intracellular CGA content up to 1.61-fold to 100.23 mg·g-1. Meanwhile, 31 primary metabolites showed significant differences, with 6 precursors related to increasing CGA biosynthesis. Secondly, the transcriptome data revealed 3637 new genes previously unannotated in the Carthamus tinctorius genome and 3653 differentially expressed genes. The genes involved in the plant signaling pathway and the biosynthesis of CGAs and their precursors showed a general up-regulation, especially the HCT gene family, which ultimately promoted CGA biosynthesis. Thirdly, the expression of a newly annotated and MeJA-responsive HCT gene (CtHCT, CtNewGene_3476) was demonstrated to be positively correlated with CGA accumulation in the cells, and transient overexpression of CtHCT enhanced CGA accumulation in tobacco. Finally, in vitro catalysis kinetics and molecular docking simulations revealed the ability and mechanism of the CtHCT protein to bind to various substrates and catalyze the formation of four hydroxycinnamic esters, including CGAs. These findings strengthened our understanding of the regulatory mechanism of CGA biosynthesis, thereby providing theoretical support for the efficient production of CGAs.

Rice responds to Spodoptera frugiperda infestation via epigenetic regulation of H3K9ac in the jasmonic acid signaling and phenylpropanoid biosynthesis pathways.

This study provided important insights into the complex epigenetic regulatory of H3K9ac-modified genes involved in the jasmonic acid signaling and phenylpropanoid biosynthesis pathways of rice in response to Spodoptera frugiperda infestation. Physiological and molecular mechanisms underlying plant responses to insect herbivores have been well studied, while epigenetic modifications such as histone acetylation and their potential regulation at the genomic level of hidden genes remain largely unknown. Histone 3 lysine 9 acetylation (H3K9ac) is an epigenetic marker widely distributed in plants that can activate gene transcription. In this study, we provided the genome-wide profiles of H3K9ac in rice (Oryza sativa) infested by fall armyworm (Spodoptera frugiperda, FAW) using CUT&Tag-seq and RNA-seq. There were 3269 and 4609 up-regulated genes identified in plants infested by FAW larvae for 3h and 12h, respectively, which were mainly enriched in alpha-linolenic acid and phenylpropanoid pathways according to transcriptomic analysis. In addition, CUT&Tag-seq analysis revealed increased H3K9ac in FAW-infested plants, and there were 422 and 543 up-regulated genes enriched with H3K9ac observed at 3h and 12h after FAW feeding, respectively. Genes with increased H3K9ac were mainly enriched in the transcription start site (TSS), suggesting that H3K9ac is related to gene transcription. Integrative analysis of both RNA-seq and CUT&Tag-seq data showed that up-expressed genes with H3K9ac enrichment were mainly involved in the jasmonic acid (JA) and phenylpropanoid pathways. Particularly, two spermidine hydroxycinnamoyl transferase genes SHT1 and SHT2 involved in phenolamide biosynthesis were highly modified by H3K9ac in FAW-infested plants. Furthermore, the Ossht1 and Ossht2 transgenic lines exhibited decreased resistance against FAW larvae. Our findings suggest that rice responds to insect herbivory via H3K9ac epigenetic regulation in the JA signaling and phenolamide biosynthesis pathways.

Effects of a multistrain Bacillus-based direct-fed microbial on gastrointestinal permeability and biomarkers of inflammation during and following feed restriction in mid-lactation Holstein cows

Objectives were to evaluate the effects of a multistrain Bacillus-based (Bacillus subtilis and Bacillus pumilus blend) direct-fed microbial (DFM) on production, metabolism, inflammation biomarkers and gastrointestinal tract (GIT) permeability during and following feed restriction (FR) in mid-lactation Holstein cows. Multiparous cows (n = 36; 138 ± 53 DIM) were randomly assigned to 1 of 3 dietary treatments: (1) control (CON; 7.5 g/d rice hulls; n = 12), (2) DFM10 (10 g/d Bacillus DFM, 4.9 × 109 cfu/d; n = 12) or 3) DFM15 (15 g/d Bacillus DFM, 7.4 × 109 cfu/d; n = 12). Before study initiation, cows were fed their respective treatments for 32 d. Cows continued to receive treatments during the trial, which consisted of 3 experimental periods (P): P1 (5 d) served as baseline for P2 (5 d), during which all cows were restricted to 40% of P1 DMI, and P3 (5 d), a "recovery" where cows were fed ad libitum. On d 4 of P1 and on d 2 and 5 of P2, GIT permeability was evaluated in vivo using the oral paracellular marker Cr-EDTA. As anticipated, FR decreased milk production, insulin, glucagon, and BUN but increased nonesterified fatty acids. During recovery, DMI rapidly increased on d 1 then subsequently decreased (4.9 kg) on d 2 before returning to baseline, whereas milk yield slowly increased but remained decreased (13%) relative to P1. The DFM10 cows had increased DMI and milk yield relative to DFM15 during P3 (10%). Overall, milk lactose content was increased in DFM cows relative to CON (0.10 percentage units), and DFM10 cows tended to have increased lactose yield relative to CON and DFM15 during P3 (8% and 10%, respectively). No overall treatment differences were observed for other milk composition variables. Circulating glucose was quadratically increased in DFM10 cows compared with CON and DFM15 during FR and recovery. Plasma Cr area under the curve was increased in all cows on d 2 (9%) and 5 (6%) relative to P1. Circulating LPS binding protein (LBP), serum amyloid A (SAA), and haptoglobin (Hp) increased in all cows during P2 compared with baseline (31%, 100%, and 9.0-fold, respectively). Circulating Hp concentrations continued to increase during P3 (274%). Overall, circulating LBP and Hp tended to be increased in DFM15 cows relative to DFM10 (29% and 81%, respectively), but no treatment differences were observed for SAA. Following feed reintroduction during P3, fecal pH initially decreased (0.62 units), but returned to baseline levels whereas fecal starch markedly increased (2.5-fold) and remained increased (82%). Absolute quantities of a fecal Butyryl-CoA CoA transferase (but) gene associated with butyrate synthesis, collected by fecal swab were increased in DFM10 cows compared with CON and DFM15 cows. In summary, FR increased GIT permeability, caused inflammation, and decreased production. Feeding DFM10 increased some key production and metabolism variables and upregulated a molecular biomarker of microbial hindgut butyrate synthesis, while DFM15 appeared to augment immune activation.

Genome-wide transcriptome profiling reveals molecular response pathways of Trichoderma harzianum in response to salt stress.

Trichoderma harzianum exhibits a strong biological control effect on many important plant pathogens, such as Fusarium oxysporum, Botrytis cinerea, and Meloidogyne. However, its biocontrol effectiveness is weakened or reduced under salt stress. The aim of this study was to investigate the molecular response of T. harzianum to salt stress at the whole-genome level. Here, we present a 44.47 Mb near-complete genome assembly of the T. harzianum qt40003 strain for the first time, which was assembled de novo with 7.59 Gb Nanopore sequencing long reads (~170-fold) and 5.2 Gb Illumina short reads (~116-fold). The assembled qt40003 genome contains 12 contigs, with a contig N50 of 4.81 Mb, in which four of the 12 contigs were entirely reconstructed in a single chromosome from telomere to telomere. The qt40003 genome contains 4.27 Mb of repeat sequences and 12,238 protein-coding genes with a BUSCO completeness of 97.5%, indicating the high accuracy and completeness of our gene annotations. Genome-wide transcriptomic analysis was used to investigate gene expression changes related to salt stress in qt40003 at 0, 2% (T2), and 4% (T4) sodium chloride concentrations. A total of 2,937 and 3,527 differentially expressed genes (DEGs) were obtained under T2 and T4 conditions, respectively. GO enrichment analysis showed that the T2-treatment DEGs were highly enriched in detoxification (p < 0.001), while the T4 DEGs were mainly enriched in cell components, mostly in cellular detoxification, cell surface, and cell wall. KEGG metabolic pathway analysis showed that 91 and 173 DEGs were significantly enriched in the T2 and T4 treatments, respectively (p < 0.01), mainly in the glutathione metabolism pathway. We further experimentally analyzed the differentially expressed glutathione transferase genes in the glutathione metabolic pathway, most of which were downregulated (13/15). In addition, we screened 13 genes related to active oxygen clearance, including six upregulated and seven downregulated genes, alongside five fungal hydrophobic proteins, of which two genes were highly expressed. Our study provides high-quality genome information for the use of T. harzianum for biological control and offers significant insights into the molecular responses of T. harzianum under salt-stress conditions.

PM2.5-induced DNA oxidative stress in A549 cells and regulating mechanisms by GST DNA methylation and Keap1/Nrf2 pathway

Fine particulate matter (PM2.5) increases the risks of lung cancer. Epigenetics provides a new toxicology mechanism for the adverse health effects of PM2.5. However, the regulating mechanisms of PM2.5 exposure on candidate gene DNA methylation changes in the development of lung cancer remain unclear. Abnormal expression of the glutathione S transferase (GST) gene is associated with cancer. However, the relationship between PM2.5 and DNA methylation-mediated GST gene expression is not well understood. In this study, we performed GST DNA methylation analysis and GST-related gene expression in human A549 cells exposed to PM2.5 (0, 50, 100 µg/mL, from Taiyuan, China) for 24 h (n = 4). We found that PM2.5 may cause DNA oxidative damage to cells and the elevation of GSTP1 promotes cell resistance to reactive oxygen species (ROS). The Kelch-1ike ECH-associated protein l (Keap1)/nuclear factor NF-E2-related factor 2 (Nrf2) pathway activates the GSTP1. The decrease in the DNA methylation level of the GSTP1 gene enhances GSTP1 expression. GST DNA methylation is associated with reduced levels of 5-methylcytosine (5mC), DNA methyltransferase 1 (DNMT1), and histone deacetylases 3 (HDAC3). The GSTM1 was not sensitive to PM2.5 stimulation. Our findings suggest that PM2.5 activates GSTP1 to defend PM2.5-induced ROS and 8-hydroxy-deoxyguanosine (8-OHdG) formation through the Keap1/Nrf2 signaling pathway and GSTP1 DNA methylation.

Common and specific genetic basis of metabolite-mediated drought responses in rice

Plants orchestrate drought responses at metabolic level but the genetic basis remains elusive in rice. In this study, 233 drought-responsive metabolites (DRMs) were quantified in a large rice population comprised of 510 diverse accessions at the reproductive stage. Large metabolic variations in drought responses were detected, and little correlation of metabolic levels between drought and normal conditions were observed. Interestingly, most of these DRMs could predict drought resistance in high accuracy. Genome-wide association study revealed 2522 significant association signals for 233 DRMs, and 98% (2471/2522) of the signals were co-localized with the association loci for drought-related phenotypic traits in the same population or the linkage-mapped QTLs for drought resistance in other populations. Totally, 10 candidate genes were efficiently identified for nine DRMs, seven of which harbored cis-eQTLs under drought condition. Based on comparative GWAS of common DRMs in rice and maize, representing irrigated and upland crops, we have identified three pairs of homologous genes associated with three DRMs between the two crops. Among the homologous genes, a transferase gene responsible for metabolic variation of N-feruloylputrescine was confirmed to confer enhanced drought resistance in rice. Our study provides not only genetic architecture of metabolic responses to drought stress in rice but also metabolic data resources to reveal the common and specific metabolite-mediated drought responses in different crops.

Genome-Wide Mapping of Quantitative Trait Loci for Yield-Attributing Traits of Peanut.

Peanuts (Arachis hypogaea L.) are important high-protein and oil-containing legume crops adapted to arid to semi-arid regions. The yield and quality of peanuts are complex quantitative traits that show high environmental influence. In this study, a recombinant inbred line population (RIL) (Valencia-C × JUG-03) was developed and phenotyped for nine traits under two environments. A genetic map was constructed using 1323 SNP markers spanning a map distance of 2003.13 cM. Quantitative trait loci (QTL) analysis using this genetic map and phenotyping data identified seventeen QTLs for nine traits. Intriguingly, a total of four QTLs, two each for 100-seed weight (HSW) and shelling percentage (SP), showed major and consistent effects, explaining 10.98% to 14.65% phenotypic variation. The major QTLs for HSW and SP harbored genes associated with seed and pod development such as the seed maturation protein-encoding gene, serine-threonine phosphatase gene, TIR-NBS-LRR gene, protein kinase superfamily gene, bHLH transcription factor-encoding gene, isopentyl transferase gene, ethylene-responsive transcription factor-encoding gene and cytochrome P450 superfamily gene. Additionally, the identification of 76 major epistatic QTLs, with PVE ranging from 11.63% to 72.61%, highlighted their significant role in determining the yield- and quality-related traits. The significant G × E interaction revealed the existence of the major role of the environment in determining the phenotype of yield-attributing traits. Notably, the seed maturation protein-coding gene in the vicinity of major QTLs for HSW can be further investigated to develop a diagnostic marker for HSW in peanut breeding. This study provides understanding of the genetic factor governing peanut traits and valuable insights for future breeding efforts aimed at improving yield and quality.

A Pinus strobus transcription factor PsbHLH1 activates the production of pinosylvin stilbenoids in transgenic Pinus koraiensis calli and tobacco leaves.

Transcription factors (TFs) play an important role in regulating the biosynthesis of secondary metabolites. In Pinus strobus, the level of methylated derivatives of pinosylvin is significantly increased upon pine wood nematode (PWN) infection, and these compounds are highly toxic to PWNs. In a previous study, we found that the expression of a basic helix-loop-helix TF gene, PsbHLH1, strongly increased in P. strobus plants after infection with PWNs. In this study, we elucidated the regulatory role of the PsbHLH1 gene in the production of methylated derivatives of pinosylvin such as pinosylvin monomethyl ether (PME) and dihydropinoylvin monomethyl ether (DPME). When PsbHLH1 was overexpressed in Pinus koraiensis calli, the production of PME and DPME was significantly increased. Overexpression of the stilbene synthase (PsSTS) and pinosylvin methyl transferase (PsPMT) genes, known as key enzymes for the biosynthesis of methylated pinosylvins, did not change PME or DPME production. Moreover, PME and DPME were not produced in tobacco leaves when the PsSTS and PsPMT genes were transiently coexpressed. However, the transient expression of three genes, PsSTS, PsPMT, and PsbHLH1, resulted in the production of PME and DPME in tobacco leaves. These results prove that PsbHLH1 is an important TF for the pinosylvin stilbene biosynthesis in pine plants and plays a regulatory role in the engineered production of PME and DPME in tobacco plants.

Detection of oxalyl-CoA decarboxylase (oxc) and formyl-CoA transferase (frc) genes in novel probiotic isolates capable of oxalate degradation in vitro

Oxalate degradation is one of lactic acid bacteria’s desirable activities. It is achieved by two enzymes, formyl coenzyme A transferase (frc) and oxalyl coenzyme A decarboxylase (oxc). The current study aimed to screen 15 locally isolated lactic acid bacteria to select those with the highest oxalate degradation ability. It also aimed to amplify the genes involved in degradation. MRS broth supplemented with 20 mM sodium oxalate was used to culture the tested isolates for 72 h. This was followed by an enzymatic assay to detect remaining oxalate. All isolates showed oxalate degradation activity to variable degrees. Five isolates demonstrated high oxalate degradation, 78 to 88%. To investigate the oxalate-degradation potential of the selected isolates, they have been further tested for the presence of genes that encode for enzymes involved in oxalate catabolism, formyl coenzyme A transferase (frc) and oxalyl coenzyme A decarboxylase (oxc). Three strains showed bands with the specific OXC and FRC forward and reverse primers designated as (SA-5, 9 and 37). Species-level identification revealed Loigolactobacillus bifermentans, Lacticaseibacillus paracasei, and Lactiplantibacillus plantarum. Preliminary results revealed that the tested probiotic strains harbored both oxc and frc whose products are putatively involved in oxalate catabolism. The probiotic potential of the selected strains was evaluated, and they showed high survival rates to both simulated gastric and intestinal fluids and variable degrees of antagonism against the tested Gram-positive and negative pathogens and were sensitive to clarithromycin but resistant to both metronidazole and ceftazidime. Finally, these strains could be exploited as an innovative approach to establish oxalate homeostasis in humans and prevent kidney stone formation.

Characterization of sugarcane mutants developed through gamma irradiations for their lignin content and caffeic acid-O-methyl transferase (COMT) gene mutations

Purpose Bagasse, the residue left after extracting juice from sugarcane stalks, is rich in lignocellulosic biomass. The lignin present in this plant biomass is the key factor that hinders the efficient extraction of ethanol from the bagasse. In the current study, γ-irradiated sugarcane mutants were evaluated for variation in lignin content and its corresponding caffeic acid-O-methyl transferase (COMT) gene. Materials and methods The acetyl bromide method was used to estimate lignin content in sugarcane mutants. PCR-based cloning of the COMT gene was performed in low lignin mutants as well as control plants in E. coli (strain DH5α) to understand the mechanism of variation at the molecular level. The Sanger sequencing for cloned gene was performed to check variation in gene sequence. Results In comparison to the control (21.5%), the mutant plants’ lignin content ranged from 13 to 28%. The Sanger sequencing revealed approximately the same length of the gene from mutants as well as a control plant. In comparison to the reference gene, the mutated gene showed SNPs and indels in different regions, which may have an impact on lignin content. Conclusions Therefore, γ-irradiated mutagenesis is an acceptable approach to develop novel mutants of sugarcane with low lignin content to enhance bioethanol production from waste material using bioprocess technology.

Genome-wide adenine N6-methylation map reveals epigenomic regulation of lipid accumulation in Nannochloropsis

Epigenetic marks on histones and DNA, such as DNA methylation at N6-adenine (6mA), play crucial roles in gene expression and genome maintenance, but their deposition and function in microalgae remain largely uncharacterized. Here, we report a genome-wide 6mA map for the model industrial oleaginous microalga Nannochloropsis oceanica produced by single-molecule real-time sequencing. Found in 0.1% of adenines, 6mA sites are mostly enriched at the AGGYV motif, more abundant in transposons and 3′ untranslated regions, and associated with active transcription. Moreover, 6mA gradually increases in abundance along the direction of gene transcription and shows special positional enrichment near splicing donor and transcription termination sites. Highly expressed genes tend to show greater 6mA abundance in the gene body than do poorly expressed genes, indicating a positive interaction between 6mA and general transcription factors. Furthermore, knockout of the putative 6mA methylase NO08G00280 by genome editing leads to changes in methylation patterns that are correlated with changes in the expression of molybdenum cofactor, sulfate transporter, glycosyl transferase, and lipase genes that underlie reductions in biomass and oil productivity. By contrast, knockout of the candidate demethylase NO06G02500 results in increased 6mA levels and reduced growth. Unraveling the epigenomic players and their roles in biomass productivity and lipid metabolism lays a foundation for epigenetic engineering of industrial microalgae.

CRISPR/Cas9-mediated Targeted Mutagenesis of Inulin Biosynthesis in Rubber Dandelion

Rubber dandelion (Taraxacum kok-saghyz) is a natural rubber-producing dandelion that has the potential to become an industrial crop. Inulin is a storage carbohydrate in rubber dandelion, and its synthesis competes with rubber production for assimilated carbon. We used the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) system to simultaneously target two sites in the gene that encodes 1-fructan:fructan-1-fructosyl transferase gene (1-FFT), a key enzyme in inulin biosynthesis. Agrobacterium rhizogenes and Agrobacterium tumefaciens-mediated plant transformation methods were used to generate transgenic plants with CRISPR/Cas9 elements. Transformation rates were 71% and 64% via A. rhizogenes and A. tumefaciens-mediated transformations, respectively. Mutagenesis was confirmed by the loss of restriction site method and Sanger sequencing. Of 13 transgenic plants obtained via A. rhizogenes, six showed editing of both target sites within the 1-FFT gene. Transgenic rubber dandelion plants were obtained within 10 weeks using A. rhizogenes-mediated transformation, which was much faster than the 6 months required for A. tumafaciens transformants. Of 11 transgenic plants obtained via A. tumefaciens, five showed mutations in both target sites. Reverse-transcription polymerase chain reaction confirmed Cas9 expression in all edited transformants. Both A. rhizogenes-mediated double-mutant transformants and A. tumefaciens-mediated double mutant transformants had lower levels of inulin than wild-type plants. Moreover, A. rhizogenes-mediated transformants had a higher rubber content than wild-type plants. Therefore, the present study validates the use of CRISPR/Cas9 gene editing as an efficient tool for the generation of useful mutations in rubber dandelion and could be implemented in future crop improvement approaches.

Assessment of tobacco (Nicotiana tabacum) and N. benthamiana as biofactories of irregular monoterpenes for sustainable crop protection

Irregular monoterpenes are important precursors of different compounds employed in pest control such as insecticides and insect sex pheromones. Metabolically engineered plants are appealing as biofactories of such compounds, but specially as potential live biodispensers of related bioactive volatiles, which could be continuously emitted to the environment from different plant tissues. Here we assess the use of cultivated tobacco and Nicotiana benthamiana as biofactories for the irregular monoterpenes chrysanthemol and lavandulol. We evaluate the impact of high levels of constitutive metabolite production on the plant physiology and biomass, and their biosynthetic dynamics for different plant tissues and developmental stages. As an example of an active pheromone compound, we super-transformed the best lavandulol-producing tobacco line with an acetyl transferase gene to obtain a tobacco lavandulyl acetate biodispenser emitting up to 0.63 mg of lavandulyl acetate per plant every day. We estimate that with these volatile emission levels, between 200 and 500 plants per hectare would be sufficient to ensure a daily emission of pheromones comparable to commercial lures. This is an important step towards plant-based sustainable solutions for pest control, and it lays the ground for further developing biofactories for other irregular monoterpenoid pheromones, whose biosynthetic genes are yet unknown.

Pharmacogenetic Effect of Thiopurine Methyl Transferase (TPMT) Gene Expression and Serum TNF on the Imuran Response in Ulcerative Colitis (UC) Iraqi Patients.

Ulcerative colitis (UC), a chronic inflammatory bowel disease (IBD), exerts its impact on both rectal and colonic mucosa, with a growing incidence. This study aims to explore the pharmacogenetic influence of thiopurine methyl transferase (TPMT) gene expression and serum tumor necrosis factor (TNF) levels on the response to Imuran in Iraqi patients with UC. Seventy individuals with chronic UC and 30 healthy controls were enrolled in this investigation. RNA extraction using the triazole method and enzyme-linked immunosorbent assay (ELISA) for TNF measurement were employed. Patients, aged 15-50 years, underwent Imuran treatment. Diverse responses to Imuran were observed among patients, with TPMT gene expression levels below 1 in 35 patients leading to side effects, while the remaining 35 patients exhibited positive responses with TPMT gene expression exceeding 1. Patients with varying degrees of severe, moderate, and mild UC associated with TNF showed a significant correlation with Imuran non-response. A distinct correlation was identified between TPMT gene expression and Imuran therapy outcomes in UC patients. Further investigation is warranted to elucidate the underlying mechanism, positioning the TPMT gene as a potential therapeutic target for mitigating the impact of UC.

Biodegradation of Benzo[a]pyrene by a White-Rot Fungus Phlebia acerina: Surfactant-Enhanced Degradation and Possible Genes Involved.

Polycyclic aromatic hydrocarbons (PAHs) are persistent environmental pollutants that pose a threat to human health. Among these PAHs, benzo[a]pyrene (BaP), a five-ring compound, exhibits high resistance to biodegradation. White-rot fungus Phlebia acerina S-LWZ20190614-6 has demonstrated higher BaP degradation capabilities compared with Phanerochaete chrysosporium and P. sordida YK-624, achieving a degradation rate of 57.7% after 32 days of incubation under a ligninolytic condition. To further enhance the biodegradation rate, three nonionic surfactants were used, and the addition of 1 or 2 g·L-1 of polyethylene glycol monododecyl ether (Brij 30) resulted in nearly complete BaP biodegradation by P. acerina S-LWZ20190614-6. Interestingly, Brij 30 did not significantly affect the activity of manganese peroxidase and lignin peroxidase, but it did decrease laccase activity. Furthermore, the impact of cytochrome P450 on BaP degradation by P. acerina S-LWZ20190614-6 was found to be relatively mild. Transcriptomic analysis provided insights into the degradation mechanism of BaP, revealing the involvement of genes related to energy production and the synthesis of active enzymes crucial for BaP degradation. The addition of Brij 30 significantly upregulated various transferase and binding protein genes in P. acerina S-LWZ20190614-6. Hence, the bioremediation potential of BaP by the white-rot fungus P. acerina S-LWZ20190614-6 holds promise and warrants further exploration.

Reciprocal regulation of TWIST1 and OGT determines the decitabine efficacy in MDS/AML

Chemoresistance poses a significant impediment to effective treatment strategies for myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Our previous study unveiled that oncogene TWIST1 interacted with DNA methyltransferase 3a (DNMT3a) to regulate the decitabine (DAC) resistance in MDS/AML. However, the underlying mechanism of TWIST1 dysregulation in DAC resistance remained enigmatic. Here, we found that O-GlcNAc modification was upregulated in CD34+ cells from MDS/AML patients who do not respond to DAC treatment. Functional study revealed that O-GlcNAcylation could stabilize TWIST1 by impeding its interaction with ubiquitin E3 ligase CBLC. In addition, as one typical transcription factor, TWIST1 could bind to the promoter of O-GlcNAc transferase (OGT) gene and activate its transcription. Collectively, we highlighted the crucial role of the O-GlcNAcylated TWIST1 in the chemoresistance capacity of MDS/AML clonal cells, which may pave the way for the development of a new therapeutic strategy targeting O-GlcNAcylated proteins and reducing the ratio of MDS/AML relapse.E9uGgDrfuE6fB_NeMXiTgkVideo

Long-term benefit of DAAs on gut dysbiosis and microbial translocation in HCV-infected patients with and without HIV coinfection

Long-term effect of Direct-acting antivirals (DAAs) on gut microbiota, short-chain fatty acids (SCFAs) and microbial translocation in patients with hepatitis C virus (HCV) infection who achieve sustained virological response (SVR) were limited. A longitudinal study of 50 patients with HCV monoinfection and 19 patients with HCV/HIV coinfection received DAAs were conducted. Fecal specimens collected at baseline and at week 72 after treatment completion (FUw72) were analyzed for 16S rRNA sequencing and the butyryl-CoA:acetateCoA transferase (BCoAT) gene expression using real-time PCR. Plasma lipopolysaccharide binding protein (LBP) and intestinal fatty acid binding protein (I-FABP) were quantified by ELISA assays. SVR rates in mono- and coinfected patients were comparable (94% vs. 100%). The improvement of gut dysbiosis and microbial translocation was found in responders but was not in non-responders. Among responders, significant restoration of alpha-diversity, BCoAT and LBP were observed in HCV patients with low-grade fibrosis (F0–F1), while HCV/HIV patients exhibited partial improvement at FUw72. I-FABP did not decline significantly in responders. Treatment induced microbiota changes with increasing abundance of SCFAs-producing bacteria, including Blautia, Fusicatenibacter, Subdoligranulum and Bifidobacterium. In conclusion, long-term effect of DAAs impacted the restoration of gut dysbiosis and microbial translocation. However, early initiation of DAAs required for an alteration of gut microbiota, enhanced SCFAs-producing bacteria, and could reduce HCV-related complications.