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SUMMARY: Plants synthesize an array of volatile compounds, many of which serve ecological roles in attracting pollinators, deterring herbivores, and communicating with their surroundings. Methyl anthranilate (MeAA) is an anti‐herbivory defensive volatile responsible for grape aroma that is emitted by several agriculturally relevant plants, including citrus, grapes, and maize. Unlike maize, which uses a one‐step anthranilate methyltransferase (AAMT), grapes have been thought to use a two‐step pathway for MeAA biosynthesis. By mining available transcriptomics data, we identified two AAMTs in Vitis vinifera (wine grape), as well as one ortholog in "Concord" grape. Many angiosperms methylate the plant hormone salicylic acid (SA) to produce methyl salicylate, which acts as a plant‐to‐plant communication molecule. Because the Citrus sinensis (sweet orange) SA methyltransferase can methylate both anthranilate (AA) and SA, we used this enzyme to examine the molecular basis of AA activity by introducing rational mutations, which identified several active site residues that increase activity with AA. Reversing this approach, we introduced mutations that imparted activity with SA in the maize AAMT, which uncovered different active site residues from those in the citrus enzyme. Sequence and phylogenetic analysis revealed that one of the Vitis AAMTs shares an ancestor with jasmonic acid methyltransferases, similar to the AAMT from strawberry (Frageria sp.). Collectively, these data demonstrate the molecular mechanisms underpinning AA activity across methyltransferases and identify one‐step enzymes by which grapes synthesize MeAA. Significance Statement: While the two‐step pathway responsible for the biosynthesis of the grape aroma molecule, methyl anthranilate, has remained incomplete in Vitis spp., we identified two one‐step anthranilate methyltransferases in wine and one in "Concord" grapes that can methylate the tryptophan pathway intermediate anthranilate. Tracing the molecular basis of anthranilate activity in the maize and sweet orange methyltransferases uncovered distinct active site amino acids that impart substrate specificity. [ABSTRACT FROM AUTHOR]

Background: SET domain-containing histone lysine methyltransferases (HKMTs) and JmjC domain-containing histone demethylases (JHDMs) are essential for maintaining dynamic changes in histone methylation across parasite development and infection. However, information on the HKMTs and JHDMs in human pathogenic piroplasms, such as Babesia duncani and Babesia microti, and in veterinary important pathogens, including Babesia bigemina, Babesia bovis, Theileria annulata and Theileria parva, is limited. Results: A total of 38 putative KMTs and eight JHDMs were identified using a comparative genomics approach. Phylogenetic analysis revealed that the putative KMTs can be divided into eight subgroups, while the JHDMs belong to the JARID subfamily, except for BdJmjC1 (BdWA1_000016) and TpJmjC1 (Tp Muguga_02g00471) which cluster with JmjC domain only subfamily members. The motifs of SET and JmjC domains are highly conserved among piroplasm species. Interspecies collinearity analysis provided insight into the evolutionary duplication events of some SET domain and JmjC domain gene families. Moreover, relative gene expression analysis by RT‒qPCR demonstrated that the putative KMT and JHDM gene families were differentially expressed in different intraerythrocytic developmental stages of B. duncani, suggesting their role in Apicomplexa parasite development. Conclusions: Our study provides a theoretical foundation and guidance for understanding the basic characteristics of several important piroplasm KMT and JHDM families and their biological roles in parasite differentiation. [ABSTRACT FROM AUTHOR]

This investigation delves into the influence of predicted microRNAs on DNA methyltransferases (DNMTs) and the PODXL gene within the NB4 cell line, aiming to elucidate their roles in the pathogenesis of acute myeloid leukemia (AML). A comprehensive methodological framework was adopted to explore the therapeutic implications of 6-gingerol on DNMTs. This encompassed a suite of bioinformatics tools for protein structure prediction, docking, molecular dynamics, and ADMET profiling, alongside empirical assessments of miRNA and PODXL expression levels. Such a multifaceted strategy facilitated an in-depth understanding of 6-gingerol’s potential efficacy in DNMT modulation. The findings indicate a nuanced interplay where 6-gingerol administration modulated miRNA expression levels, decreasing in DNMT1 and DNMT3A expression in NB4 cells. This alteration indirectly influenced PODXL expression, contributing to the manifestation of oncogenic phenotypes. The overexpression of DNMT1 and DNMT3A in NB4 cells may contribute to AML, which appears modulable via microRNAs such as miR-193a and miR-200c. Post-treatment with 6-gingerol, DNMT1 and DNMT3A expression alterations were observed, culminating in the upregulation of miR-193a and miR-200c. This cascade effect led to the dysregulation of tumor suppressor genes in cancer cells, including downregulation of PODXL, and the emergence of cancerous traits. These insights underscore the therapeutic promise of 6-gingerol in targeting DNMTs and microRNAs within the AML context. [ABSTRACT FROM AUTHOR]

Methylation of RNA nucleotides represents an important layer of gene expression regulation, and perturbation of the RNA methylome is associated with pathophysiology. In cells, RNA methylations are installed by RNA methyltransferases (RNMTs) that are specialized to catalyze particular types of methylation (ribose or different base positions). Furthermore, RNMTs must specifically recognize their appropriate target RNAs within the RNA-dense cellular environment. Some RNMTs are catalytically active alone and achieve target specificity via recognition of sequence motifs and/or RNA structures. Others function together with protein cofactors that can influence stability, S-adenosyl-L-methionine binding, and RNA affinity as well as aiding specific recruitment and catalytic activity. Association of RNMTs with guide RNAs represents an alternative mechanism to direct site-specific methylation by an RNMT that lacks intrinsic specificity. Recently, ribozyme-catalyzed methylation of RNA has been achieved in vitro, and here, we compare these different strategies for RNA methylation from structural and mechanistic perspectives. [ABSTRACT FROM AUTHOR]

Introduction: The African oil palm (Elaeis guineensis Jacq.) is the predominant oil crop in the world. In addition to triacylglycerols, crude palm oil (CPO) extracted from the mesocarp of the fruits, contains high amounts of provitamin A (carotenes) and vitamin E (tocochromanols). Because of their unsaturated nature, the carotenes are prone to oxidation and therefore are in part limiting for the shelf life of CPO. Results: In the present study, a wild Oil Palm tree (C59) from Cameroon was identified that lacks α-tocopherol and α-tocotrienol and instead accumulates the respective γ forms, suggesting that the activity of γ-tocopherol methyltransferase (VTE4) was affected. Sequencing of the VTE4 locus in the genome of plant C59 identified a G/C polymorphism that causes the exchange of a highly conserved tryptophan at position 290 with serine. The W290S exchange renders the VTE4 enzyme inactive, as shown after expression in Escherichia coli and Arabidopsis thaliana. The oxidative stability of carotenes in the mesocarp of the wild palm C59 was enhanced compared with control accessions. Furthermore, supplementation of commercial palm oil with different tocochromanols showed that γ-tocotrienol exerts a stronger effect during the protection of carotenes against oxidation than α-tocotrienol. Methods: A tree with unusual toochromanol composition was identified by HPLC screening of the mesocarp of wild trees. Polymorphisms in a candidate gene were identified by DNA sequencing. The candidate protein was heterologously expressed in Escherichia coli coli and Arabidopsis thaliana to test for enzyme activity. Oxidative stability of the CPO was studied by following carotene degradation over time. Discussion: Therefore, the introduction of the high γ-tocotrienol trait into elite breeding lines represents a potent strategy to protect carotenes against oxidation and extend the shelf life of CPO, hence allowing the development of a value added high-carotene CPO to be used to fight against vitamin A deficiency [ABSTRACT FROM AUTHOR]

DNA methylation is an important epigenetic regulatory mechanism, it regulates gene expression by recruiting proteins involved in gene repression or by inhibiting the binding of transcription factor(s) to DNA. In this study, a novel methyltransferase 2a gene (Zmet2a) was cloned in maize and identified by polymerase chain reaction-base (PCR-base) using a bioinformatics strategy. The Zmet2a cDNA sequence is 2739 bp long and translates to 912 amino acid peptides. The Zmet2a protein revealed that it contains BAH and CHROMO structural domains, is a non-transmembrane protein that is hydrophilically unstable, and has no signal peptide structure. Meanwhile, we verified the biological roles of Zmet2a using transgenic Arabidopsis overexpressing Zmet2a and Zmet2a -knockout maize. Transgenic Zmet2a Arabidopsis thaliana showed highly significant advancement in flowering time, and Zmet2a -knockout maize showed advancement in flowering time, with significant changes in several traits. Altogether, these report the role of Zmet2a in the regulation of flowering time, which will lay a foundation for revealing the biological function and epigenetic regulation mechanism of Zmet2a in the growth, development and flowering of maize. Supplementary Material Supplementary Material File [ABSTRACT FROM AUTHOR]

The immune system comprises a complex yet tightly regulated network of cells and molecules that play a critical role in protecting the body from infection and disease. The activity and development of each immune cell is regulated in a myriad of ways including through the cytokine milieu, the availability of key receptors, via tailored intracellular signalling cascades, dedicated transcription factors and even by directly modulating gene accessibility and expression; the latter is more commonly known as epigenetic regulation. In recent years, epigenetic regulators have begun to emerge as key players involved in modulating the immune system. Among these, the lysine methyltransferase DOT1L has gained significant attention for its involvement in orchestrating immune cell formation and function. In this review we provide an overview of the role of DOT1L across the immune system and the implications of this role on health and disease. We begin by elucidating the general mechanisms of DOT1Lmediated histone methylation and its impact on gene expression within immune cells. Subsequently, we provide a detailed and comprehensive overview of recent studies that identify DOT1L as a crucial regulator of immune cell development, differentiation, and activation. Next, we discuss the potential mechanisms of DOT1L-mediated regulation of immune cell function and shed light on how DOT1L might be contributing to immune cell homeostasis and dysfunction. We then provide food for thought by highlighting some of the current obstacles and technical limitations precluding a more in-depth elucidation of DOT1L's role. Finally, we explore the potential therapeutic implications of targeting DOT1L in the context of immune-related diseases and discuss ongoing research efforts to this end. Overall, this review consolidates the current paradigm regarding DOT1L's role across the immune network and emphasises its critical role in governing the healthy immune system and its potential as a novel therapeutic target for immune-related diseases. A deeper understanding of DOT1L's immunomodulatory functions could pave the way for innovative therapeutic approaches which fine-tune the immune response to enhance or restore human health. [ABSTRACT FROM AUTHOR]

Abstract Background SET domain-containing histone lysine methyltransferases (HKMTs) and JmjC domain-containing histone demethylases (JHDMs) are essential for maintaining dynamic changes in histone methylation across parasite development and infection. However, information on the HKMTs and JHDMs in human pathogenic piroplasms, such as Babesia duncani and Babesia microti, and in veterinary important pathogens, including Babesia bigemina, Babesia bovis, Theileria annulata and Theileria parva, is limited. Results A total of 38 putative KMTs and eight JHDMs were identified using a comparative genomics approach. Phylogenetic analysis revealed that the putative KMTs can be divided into eight subgroups, while the JHDMs belong to the JARID subfamily, except for BdJmjC1 (BdWA1_000016) and TpJmjC1 (Tp Muguga_02g00471) which cluster with JmjC domain only subfamily members. The motifs of SET and JmjC domains are highly conserved among piroplasm species. Interspecies collinearity analysis provided insight into the evolutionary duplication events of some SET domain and JmjC domain gene families. Moreover, relative gene expression analysis by RT‒qPCR demonstrated that the putative KMT and JHDM gene families were differentially expressed in different intraerythrocytic developmental stages of B. duncani, suggesting their role in Apicomplexa parasite development. Conclusions Our study provides a theoretical foundation and guidance for understanding the basic characteristics of several important piroplasm KMT and JHDM families and their biological roles in parasite differentiation.

Abstract This investigation delves into the influence of predicted microRNAs on DNA methyltransferases (DNMTs) and the PODXL gene within the NB4 cell line, aiming to elucidate their roles in the pathogenesis of acute myeloid leukemia (AML). A comprehensive methodological framework was adopted to explore the therapeutic implications of 6-gingerol on DNMTs. This encompassed a suite of bioinformatics tools for protein structure prediction, docking, molecular dynamics, and ADMET profiling, alongside empirical assessments of miRNA and PODXL expression levels. Such a multifaceted strategy facilitated an in-depth understanding of 6-gingerol’s potential efficacy in DNMT modulation. The findings indicate a nuanced interplay where 6-gingerol administration modulated miRNA expression levels, decreasing in DNMT1 and DNMT3A expression in NB4 cells. This alteration indirectly influenced PODXL expression, contributing to the manifestation of oncogenic phenotypes. The overexpression of DNMT1 and DNMT3A in NB4 cells may contribute to AML, which appears modulable via microRNAs such as miR-193a and miR-200c. Post-treatment with 6-gingerol, DNMT1 and DNMT3A expression alterations were observed, culminating in the upregulation of miR-193a and miR-200c. This cascade effect led to the dysregulation of tumor suppressor genes in cancer cells, including downregulation of PODXL, and the emergence of cancerous traits. These insights underscore the therapeutic promise of 6-gingerol in targeting DNMTs and microRNAs within the AML context.

FAM86A is a class I lysine methyltransferase (KMT) that generates trimethylation on the eukaryotic translation elongation factor 2 (EEF2) at Lys525. Publicly available data from The Cancer Dependency Map project indicate high dependence of hundreds of human cancer cell lines on FAM86A expression. This classifies FAM86A among numerous other KMTs as potential targets for future anticancer therapies. However, selective inhibition of KMTs by small molecules can be challenging due to high conservation within the S-adenosyl methionine (SAM) cofactor binding domain among KMT subfamilies. Therefore, understanding the unique interactions within each KMT-substrate pair can facilitate developing highly specific inhibitors. The FAM86A gene encodes an N-terminal FAM86 domain of unknown function in addition to its C-terminal methyltransferase domain. Here, we used a combination of X-ray crystallography, the AlphaFold algorithms, and experimental biochemistry to identify an essential role of the FAM86 domain in mediating EEF2 methylation by FAM86A. To facilitate our studies, we also generated a selective EEF2K525 methyl antibody. Overall, this is the first report of a biological function for the FAM86 structural domain in any species and an example of a noncatalytic domain participating in protein lysine methylation. The interaction between the FAM86 domain and EEF2 provides a new strategy for developing a specific FAM86A small molecule inhibitor, and our results provide an example in which modeling a protein-protein interaction with AlphaFold expedites experimental biology.

Ribosomes are large macromolecular complexes composed of both proteins and RNA, that require a plethora of factors and post-transcriptional modifications for their biogenesis. In human mitochondria, the ribosomal RNA is post-transcriptionally modified at ten sites. The N4-methylcytidine (m4C) methyltransferase, METTL15, modifies the 12S rRNA of the small subunit at position C1486. The enzyme is essential for mitochondrial protein synthesis and assembly of the mitoribosome small subunit, as shown here and by previous studies. Here, we demonstrate that the m4C modification is not required for small subunit biogenesis, indicating that the chaperone-like activity of the METTL15 protein itself is an essential component for mitoribosome biogenesis.

Abstract Background This study explored the impact of predicted miRNAs on DNA methyltransferases (DNMTs) and the PODXL gene in Nalm6 cells, revealing the significance of these miRNAs in acute lymphocytic leukemia (ALL). Methods A comprehensive approach was adopted, integrating bioinformatic analyses encompassing protein structure prediction, molecular docking, dynamics, and ADMET profiling, in conjunction with evaluations of gene and miRNA expression patterns. This methodology was employed to elucidate the therapeutic potential of catechin compounds in modulating the activity of DNA methyltransferases (DNMTs) and the PODXL gene. Results The findings from our investigation indicate that catechins possess the capability to inhibit DNMT enzymes. This inhibitory effect is associated with the upregulation of microRNAs miR-200c and miR-548 and a concurrent downregulation of PODXL gene expression. These molecular interactions culminate in an augmented apoptotic response within ALL (Nalm6) cells. Conclusion The study posits that catechins may represent a viable therapeutic avenue for inducing apoptosis in ALL cells. This is achieved through the modulation of epigenetic mechanisms and alterations in gene expression profiles, highlighting the potential of catechins as agents for cancer therapy.

Abstract Background The adult intestinal epithelium is a complex, self-renewing tissue composed of specialized cell types with diverse functions. Intestinal stem cells (ISCs) located at the bottom of crypts, where they divide to either self-renew, or move to the transit amplifying zone to divide and differentiate into absorptive and secretory cells as they move along the crypt-villus axis. Enteroendocrine cells (EECs), one type of secretory cells, are the most abundant hormone-producing cells in mammals and involved in the control of energy homeostasis. However, regulation of EEC development and homeostasis is still unclear or controversial. We have previously shown that protein arginine methyltransferase (PRMT) 1, a histone methyltransferase and transcription co-activator, is important for adult intestinal epithelial homeostasis. Results To investigate how PRMT1 affects adult intestinal epithelial homeostasis, we performed RNA-Seq on small intestinal crypts of tamoxifen-induced intestinal epithelium-specific PRMT1 knockout and PRMT1fl/fl adult mice. We found that PRMT1fl/fl and PRMT1-deficient small intestinal crypts exhibited markedly different mRNA profiles. Surprisingly, GO terms and KEGG pathway analyses showed that the topmost significantly enriched pathways among the genes upregulated in PRMT1 knockout crypts were associated with EECs. In particular, genes encoding enteroendocrine-specific hormones and transcription factors were upregulated in PRMT1-deficient small intestine. Moreover, a marked increase in the number of EECs was found in the PRMT1 knockout small intestine. Concomitantly, Neurogenin 3-positive enteroendocrine progenitor cells was also increased in the small intestinal crypts of the knockout mice, accompanied by the upregulation of the expression levels of downstream targets of Neurogenin 3, including Neuod1, Pax4, Insm1, in PRMT1-deficient crypts. Conclusions Our finding for the first time revealed that the epigenetic enzyme PRMT1 controls mouse enteroendocrine cell development, most likely via inhibition of Neurogenin 3-mediated commitment to EEC lineage. It further suggests a potential role of PRMT1 as a critical transcriptional cofactor in EECs specification and homeostasis to affect metabolism and metabolic diseases.

Abstract Background Syringic acid (SA) is a high-value natural compound with diverse biological activities and wide applications, commonly found in fruits, vegetables, and herbs. SA is primarily produced through chemical synthesis, nonetheless, these chemical methods have many drawbacks, such as considerable equipment requirements, harsh reaction conditions, expensive catalysts, and numerous by-products. Therefore, in this study, a novel biotransformation route for SA production was designed and developed by using engineered whole cells. Results An O-methyltransferase from Desulfuromonas acetoxidans (DesAOMT), which preferentially catalyzes a methyl transfer reaction on the meta-hydroxyl group of catechol analogues, was identified. The whole cells expressing DesAOMT can transform gallic acid (GA) into SA when S-adenosyl methionine (SAM) is used as a methyl donor. We constructed a multi-enzyme cascade reaction in Escherichia coli, containing an endogenous shikimate kinase (AroL) and a chorismate lyase (UbiC), along with a p-hydroxybenzoate hydroxylase mutant (PobA**) from Pseudomonas fluorescens, and DesAOMT; SA was biosynthesized from shikimic acid (SHA) by using whole cells catalysis. The metabolic system of chassis cells also affected the efficiency of SA biosynthesis, blocking the chorismate metabolism pathway improved SA production. When the supply of the cofactor NADPH was optimized, the titer of SA reached 133 μM (26.2 mg/L). Conclusion Overall, we designed a multi-enzyme cascade in E. coli for SA biosynthesis by using resting or growing whole cells. This work identified an O-methyltransferase (DesAOMT), which can catalyze the methylation of GA to produce SA. The multi-enzyme cascade containing four enzymes expressed in an engineered E. coli for synthesizing of SA from SHA. The metabolic system of the strain and biotransformation conditions influenced catalytic efficiency. This study provides a new green route for SA biosynthesis. Graphical Abstract

Hangsheng Zhou,1,2,* Jiandong Gui,1,2,* Lijie Zhu,2 Yuanyuan Mi2 1Wuxi Medical College, Jiangnan University, Wuxi, Jiangsu Province, 214122, People’s Republic of China; 2Department of Urology, Affiliated Hospital of Jiangnan University, Wuxi, Jiangsu Province, 214122, People’s Republic of China*These authors contributed equally to this workCorrespondence: Yuanyuan Mi; Lijie Zhu, Department of Urology, Affiliated Hospital of Jiangnan University, No. 1000 Hefeng Road, Wuxi, Jiangsu, 214122, People’s Republic of China, Email miniao1984@163.com; jndxfyzlj@163.comAbstract: Methylation-mediated gene silencing is closely related to the occurrence and development of human tumors. The euchromatic histone lysine methyltransferase 2 (EHMT2, also known as G9a) is highly expressed in many tumors and is generally considered to be an oncogene, which is associated with the poor outcome of many tumors. Combined immunotherapy and immune checkpoint blockade therapy also have good efficacy and certain safety. However, there are still many difficulties in the drugs targeting G9a, and the combined effect and safety of G9a with many drugs is still under study. This article aims to summarize the role and mechanism of G9a and its inhibitors in tumors in the past two years, and to understand the application prospect of G9a from the perspective of diagnosis and treatment.Keywords: cancer, G9a, methyltransferase, function, mechanism

In the contemporary epoch, cancer stands as the predominant cause of premature global mortality, necessitating a focused exploration of molecular markers and advanced therapeutic strategies. N6-methyladenosine (m6A), the most prevalent mRNA modification, undergoes dynamic regulation by enzymes referred to as methyltransferases (writers), demethylases (erasers), and effective proteins (readers). Despite lacking methylation activity, RNA-binding motif protein 15 (RBM15), a member of the m6A writer family, assumes a crucial role in recruiting the methyltransferase complex (MTC) and binding to mRNA. Although the impact of m6A modifications on cancer has garnered widespread attention, RBM15 has been relatively overlooked. This review briefly outlines the structure and operational mechanism, and delineates the unique role of RBM15 in various cancers, shedding light on its molecular basis and providing a groundwork for potential tumor-targeted therapies. [ABSTRACT FROM AUTHOR]

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and irreversible interstitial lung disease with a prognosis worse than lung cancer. It is a fatal lung disease with largely unknown etiology and pathogenesis, and no effective therapeutic drugs render its treatment largely unsuccessful. With continuous indepth research efforts, the epigenetic mechanisms in IPF pathogenesis have been further discovered and concerned. As a widely studied mechanism of epigenetic modification, DNA methylation is primarily facilitated by DNA methyltransferases (DNMTs), resulting in the addition of a methyl group to the fifth carbon position of the cytosine base, leading to the formation of 5-methylcytosine (5-mC). Dysregulation of DNA methylation is intricately associated with the advancement of respiratory disorders. Recently, the role of DNA methylation in IPF pathogenesis has also received considerable attention. DNA methylation patterns include methylation modification and demethylation modification and regulate a range of essential biological functions through gene expression regulation. The Ten-Eleven-Translocation (TET) family of DNA dioxygenases is crucial in facilitating active DNA demethylation through the enzymatic conversion of the modified genomic base 5-mC to 5-hydroxymethylcytosine (5-hmC). TET2, a member of TET proteins, is involved in lung inflammation, and its protein expression is downregulated in the lungs and alveolar epithelial type II cells of IPF patients. This review summarizes the current knowledge of pathologic features and DNA methylation mechanisms of pulmonary fibrosis, focusing on the critical roles of abnormal DNA methylation patterns, DNMTs, and TET proteins in impacting IPF pathogenesis. Researching DNA methylation will enchance comprehension of the fundamental mechanisms involved in IPF pathology and provide novel diagnostic biomarkers and therapeutic targets for pulmonary fibrosis based on the studies involving epigenetic mechanisms. [ABSTRACT FROM AUTHOR]

Objective To observe the effect of methyltransferase SETDB1 on the migration and invasion of oral squamous cell carcinoma cells, and to explore the related mechanisms. Methods Different concentrations of methyltransferase inhibitors GSK3685032 (0，0. 2，0. 4，0. 6，0. 8 µmol/L) acted on the oral squamous cell carcinoma CAL-27 cells. Cell proliferative viability was measured by the MTT method, and we found that the optimal action concentration of GSK3685032 was 0. 6 µmol/L. CAL-27 cells were divided into the control and experimental groups; cells in the control group were treated with organic solvent DMSO, and cells in the experimental group were treated with 0. 6 µmol/L GSK3685032. Cell SETDB1 mRNA was measured by RT-qPCR, and the SETDB1 protein was detected by Western blotting. Cell migration ability was detected by Scratch assay, the invasion ability was detected by Transwell chamber invasion assay, and SOX 7 methylation level was tested by pyrosequencing. Results The mRNA and protein expression levels of SETDB1 were lower in the experimental group than in the control group (both P<0. 05). The migration distance of the experimental group was less than that of the control group, and the number of transmenbrance cells was smaller than that of the control group (P<0. 05). The methylation rate of SOX 7 in the experimental group was lower than that of the control group (P<0. 05). Conclusion Methyltransferase SETDB1 can inhibit the migration and invasion of oral squamous cell carcinoma cells, and the mechanism may be related to the down-regulation of intracellular SOX7 methylation level. [ABSTRACT FROM AUTHOR]

Background: This study explored the impact of predicted miRNAs on DNA methyltransferases (DNMTs) and the PODXL gene in Nalm6 cells, revealing the significance of these miRNAs in acute lymphocytic leukemia (ALL). Methods: A comprehensive approach was adopted, integrating bioinformatic analyses encompassing protein structure prediction, molecular docking, dynamics, and ADMET profiling, in conjunction with evaluations of gene and miRNA expression patterns. This methodology was employed to elucidate the therapeutic potential of catechin compounds in modulating the activity of DNA methyltransferases (DNMTs) and the PODXL gene. Results: The findings from our investigation indicate that catechins possess the capability to inhibit DNMT enzymes. This inhibitory effect is associated with the upregulation of microRNAs miR-200c and miR-548 and a concurrent downregulation of PODXL gene expression. These molecular interactions culminate in an augmented apoptotic response within ALL (Nalm6) cells. Conclusion: The study posits that catechins may represent a viable therapeutic avenue for inducing apoptosis in ALL cells. This is achieved through the modulation of epigenetic mechanisms and alterations in gene expression profiles, highlighting the potential of catechins as agents for cancer therapy. [ABSTRACT FROM AUTHOR]

Indoles and pyrroloindoles are structural motifs present in many biologically active natural products. Multiple classes of enzymes catalyze the transformation of indoles into pyrroloindoles via group transfer followed by intramolecular cyclization, such as peroxydases, methyltransferases, and prenyltransferases. Due to the selective introduction of a stereogenic center, these enzymes receive increasing attention as catalytic tools for the production of pharmacologically relevant compounds. Two new colorimetric assays are described in this work, which allow for the quantification of such enzymatic reactions from the perspective of the substrate and the product. For the substrates, the indole assay is based on a modified version of the Ehrlich test, with the use of light as a driving force for color formation. The pyrroloindole assay uses cerium sulfate as a reagent for the colorimetric quantification of the enzymatic products. The assays are complementary and both were successfully utilized for enzymatic activity determination of a C3‐indole methyltransferase. They can facilitate high‐throughput screening of mutant libraries, offering support for the engineering of such enzymes, but can also be used as stand‐alone methods for the detection and quantification of natural products. [ABSTRACT FROM AUTHOR]

Background: The adult intestinal epithelium is a complex, self-renewing tissue composed of specialized cell types with diverse functions. Intestinal stem cells (ISCs) located at the bottom of crypts, where they divide to either self-renew, or move to the transit amplifying zone to divide and differentiate into absorptive and secretory cells as they move along the crypt-villus axis. Enteroendocrine cells (EECs), one type of secretory cells, are the most abundant hormone-producing cells in mammals and involved in the control of energy homeostasis. However, regulation of EEC development and homeostasis is still unclear or controversial. We have previously shown that protein arginine methyltransferase (PRMT) 1, a histone methyltransferase and transcription co-activator, is important for adult intestinal epithelial homeostasis. Results: To investigate how PRMT1 affects adult intestinal epithelial homeostasis, we performed RNA-Seq on small intestinal crypts of tamoxifen-induced intestinal epithelium-specific PRMT1 knockout and PRMT1fl/fl adult mice. We found that PRMT1fl/fl and PRMT1-deficient small intestinal crypts exhibited markedly different mRNA profiles. Surprisingly, GO terms and KEGG pathway analyses showed that the topmost significantly enriched pathways among the genes upregulated in PRMT1 knockout crypts were associated with EECs. In particular, genes encoding enteroendocrine-specific hormones and transcription factors were upregulated in PRMT1-deficient small intestine. Moreover, a marked increase in the number of EECs was found in the PRMT1 knockout small intestine. Concomitantly, Neurogenin 3-positive enteroendocrine progenitor cells was also increased in the small intestinal crypts of the knockout mice, accompanied by the upregulation of the expression levels of downstream targets of Neurogenin 3, including Neuod1, Pax4, Insm1, in PRMT1-deficient crypts. Conclusions: Our finding for the first time revealed that the epigenetic enzyme PRMT1 controls mouse enteroendocrine cell development, most likely via inhibition of Neurogenin 3-mediated commitment to EEC lineage. It further suggests a potential role of PRMT1 as a critical transcriptional cofactor in EECs specification and homeostasis to affect metabolism and metabolic diseases. [ABSTRACT FROM AUTHOR]

Primary antibodies are one of the main tools used in molecular biology research. However, the often-occurring cross-reactivity of primary antibodies complicates accurate data analysis. Our results show that three commercial polyclonal antibodies raised against N-6 adenine-specific DNA methyltransferase 1 (N6AMT1) strongly cross-react with endogenous and recombinant mitosis-associated protein Aurora kinase A (AURKA). The cross-reactivity was verified through immunofluorescence, immunoblot, and immunoprecipitation assays combined with mass spectrometry. N6AMT1 and AURKA are evolutionarily conserved proteins that are vital for cellular processes. Both proteins share the motif ENNPEE, which is unique to only these two proteins. We suggest that N6AMT1 antibodies recognise this motif in N6AMT1 and AURKA proteins and exhibit an example of "specific" non-specificity. This serves as an example of the importance of controls and critical data interpretation in molecular biology research. [ABSTRACT FROM AUTHOR]

Caffeine biosynthesis in Coffea evolved independently through tandem gene duplication and subsequent neofunctionalization of ancestral N-methyltransferases (NMTs). The bulk of our understanding of this process in Coffea is centered on two species: C. arabica and C. canephora. To gain a wider perspective of caffeine biosynthesis in Coffea, this study focused on the less popular but commercially available C. liberica var. liberica, locally known as "kapeng barako." This study aimed to determine the genomic and coding sequences of the three NMTs -- mainly, xanthosine methyltransferase (XMT), 7-methylxanthine-N-methyltransferase (MXMT), and dimethylxanthine methyltransferase (DXMT) -- through Sanger sequencing. Relative expression levels of these NMTs in the leaves, flower bud, green fruit, and red fruit stages were also determined through polymerase chain reaction amplification of synthesized cDNA and then gel electrophoresis. Results reveal that the genetic sequences of NMTs of C. liberica were similar to the C. eugenioides-derived NMTs of C. arabica. Results also indicate that there is a possibility that more than one NMT gene for each NMT cluster (XMT, MXMT, DXMT) exists. This study has also identified genomic sequences and transcripts, whose sequences were in between the XMT and MXMT gene clusters. The function of these novel sequences is yet to be determined. NMT expression levels in C. liberica were similar to those observed in C. arabica and C. canephora, wherein expression peaks at the green fruit stage and then diminishes, as the fruit ripens fully. Although the NMTs of C. liberica have a lot of similarities with C. arabica and C. canephora, discovering the function and role in caffeine biosynthesis of these novel genomic sequences and transcripts will help us further understand the evolution of caffeine biosynthesis in Coffea. [ABSTRACT FROM AUTHOR]

Objectives To characterize and elucidate the spread of amikacin-resistant Enterobacteriaceae isolates from environmental samples on a pig farm in the UK, following the previous identification of index Salmonella isolates harbouring the rmtB gene, a 16S rRNA methylase. Methods Environmental samples were collected during two visits to a pig farm in the UK. Isolates were recovered using selective media (amikacin 128 mg/L) followed by real-time PCR and WGS to analyse rmtB -carrying Salmonella and Escherichia coli isolates. Results Salmonella and E. coli isolates harbouring the rmtB gene were detected at both farm visits. All Salmonella isolates were found to be monophasic S. enterica serovar Typhimurium variant Copenhagen of ST34. rmtB -harbouring E. coli isolates were found to be one of three STs: ST4089, ST1684 and ST34. Long-read sequencing identified the rmtB gene to be chromosomally located in Salmonella isolates and on IncFII-type plasmids in E. coli isolates. The results showed the rmtB gene to be flanked by IS 26 elements and several resistance genes. Conclusions We report on the occurrence of rmtB -harbouring Enterobacteriaceae on a pig farm in the UK. rmtB confers resistance to multiple aminoglycosides and this work highlights the need for surveillance to assess dissemination and risk. [ABSTRACT FROM AUTHOR]

Grass xylan consists of a linear chain of β-1,4-linked xylosyl residues that often form domains substituted only with either arabinofuranose (Ara f) or glucuronic acid (GlcA)/methylglucuronic acid (MeGlcA) residues, and it lacks the unique reducing end tetrasaccharide sequence found in dicot xylan. The mechanism of how grass xylan backbone elongation is initiated and how its distinctive substitution pattern is determined remains elusive. Here, we performed biochemical characterization of rice xylan biosynthetic enzymes, including xylan synthases, glucuronyltransferases and methyltransferases. Activity assays of rice xylan synthases demonstrated that they required short xylooligomers as acceptors for their activities. While rice xylan glucuronyltransferases effectively glucuronidated unsubstituted xylohexaose acceptors, they transferred little GlcA residues onto (Ara f)-substituted xylohexaoses and rice xylan 3- O -arabinosyltransferase could not arabinosylate GlcA-substituted xylohexaoses, indicating that their intrinsic biochemical properties may contribute to the distinctive substitution patterns of rice xylan. In addition, we found that rice xylan methyltransferase exhibited a low substrate binding affinity, which may explain the partial GlcA methylation in rice xylan. Furthermore, immunolocalization of xylan in xylem cells of both rice and Arabidopsis showed that it was deposited together with cellulose in secondary walls without forming xylan-rich nanodomains. Together, our findings provide new insights into the biochemical mechanisms underlying xylan backbone elongation and substitutions in grass species. [ABSTRACT FROM AUTHOR]

Ribosomal RNAs (rRNAs) are extensively modified during the transcription and subsequent maturation. Three types of modifications, 2′-O-methylation of ribose moiety, pseudouridylation, and base modifications, are introduced either by a snoRNA-driven mechanism or by stand-alone enzymes. Modified nucleotides are clustered at the functionally important sites, including peptidyl transferase center (PTC). Therefore, it has been hypothesised that the modified nucleotides play an important role in ensuring the functionality of the ribosome. In this study, we demonstrate that seven 25S rRNA modifications, including four evolutionarily conserved modifications, in the proximity of PTC can be simultaneously depleted without loss of cell viability. Yeast mutants lacking three snoRNA genes (snR34, snR52, and snR65) and/or expressing enzymatically inactive variants of spb1(D52A/E679K) and nop2(C424A/C478A) were constructed. The results show that rRNA modifications in PTC contribute collectively to efficient translation in eukaryotic cells. The deficiency of seven modified nucleotides in 25S rRNA resulted in reduced cell growth, cold sensitivity, decreased translation levels, and hyperaccurate translation, as indicated by the reduced missense and nonsense suppression. The modification m5C2870 is crucial in the absence of the other six modified nucleotides. Thus, the pattern of rRNA-modified nucleotides around the PTC is essential for optimal ribosomal translational activity and translational fidelity. [ABSTRACT FROM AUTHOR]

As the most abundant form of methylation modification in messenger RNA (mRNA), the distribution of N6‐methyladenosine (m6A) has been preliminarily revealed in herbaceous plants under salt stress, but its function and mechanism in woody plants were still unknown. Here, we showed that global m6A levels increased during poplar response to salt stress. Methylated RNA immunoprecipitation sequencing (MeRIP‐seq) revealed that m6A significantly enriched in the coding sequence region and 3′‐untranslated regions in poplar, by recognising the conserved motifs, AGACU, GGACA and UGUAG. A large number of differential m6A transcripts have been identified, and some have been proved involving in salt response and plant growth and development. Further combined analysis of MeRIP‐seq and RNA‐seq revealed that the m6A hypermethylated and enrich in the CDS region preferred to positively regulate expression abundance. Writer inhibitor, 3‐deazaneplanocin A treatment increased the sensitivity of poplar to salt stress by reducing mRNA stability to regulate the expression of salt‐responsive transcripts PagMYB48, PagGT2, PagNAC2, PagGPX8 and PagARF2. Furthermore, we verified that the methyltransferase PagFIP37 plays a positively role in the response of poplar to salt stress, overexpressed lines have stronger salt tolerance, while RNAi lines were more sensitive to salt, which relied on regulating mRNA stability in an m6A manner of salt‐responsive transcripts PagMYB48, PagGT2, PagNAC2, PagGPX8 and PagARF2. Collectively, these results revealed the regulatory role of m6A methylation in poplar response to salt stress, and revealed the importance and mechanism of m6A methylation in the response of woody plants to salt stress for the first time. Summary statement: We revealed the regulatory role of m6A methylation in poplar response to salt stress, and revealed the importance and mechanism of m6A methylation in the response of woody plants to salt stress for the first time. [ABSTRACT FROM AUTHOR]

Main conclusion: Six methyltransferase genes affecting tomato fruit ripening were identified through genome-wide screening, VIGS assay, and expression pattern analysis. The data provide the basis for understanding new mechanisms of methyltransferases. Fruit ripening is a critical stage for the formation of edible quality and seed maturation, which is finely modulated by kinds of factors, including genetic regulators, hormones, external signals, etc. Methyltransferases (MTases), important genetic regulators, play vital roles in plant development through epigenetic regulation, post-translational modification, or other mechanisms. However, the regulatory functions of numerous MTases except DNA methylation in fruit ripening remain limited so far. Here, six MTases, which act on different types of substrates, were identified to affect tomato fruit ripening. First, 35 MTase genes with relatively high expression at breaker (Br) stage of tomato fruit were screened from the tomato MTase gene database encompassing 421 genes totally. Thereafter, six MTase genes were identified as potential regulators of fruit ripening via virus-induced gene silencing (VIGS), including four genes with a positive regulatory role and two genes with a negative regulatory role, respectively. The expression of these six MTase genes exhibited diverse patterns during the fruit ripening process, and responded to various external ripening-related factors, including ethylene, 1-methylcyclopropene (1-MCP), temperature, and light exposure. These results help to further elaborate the biological mechanisms of MTase genes in tomato fruit ripening and enrich the understanding of the regulatory mechanisms of fruit ripening involving MTases, despite of DNA MTases. [ABSTRACT FROM AUTHOR]

As(III) S‐adenosylmethionine methyltransferases, pivotal enzymes in arsenic metabolism, facilitate the methylation of arsenic up to three times. This process predominantly yields trivalent mono‐ and dimethylarsenite, with trimethylarsine forming in smaller amounts. While this enzyme acts as a detoxifier in microbial systems by altering As(III), in humans, it paradoxically generates more toxic and potentially carcinogenic methylated arsenic species. The strong affinity of As(III) for cysteine residues, forming As(III)‐thiolate bonds, is exploited in medical treatments, notably in arsenic trioxide (Trisenox®), an FDA‐approved drug for leukemia. The effectiveness of this drug is partly due to its interaction with cysteine residues, leading to the breakdown of key oncogenic fusion proteins. In this study, we extend the understanding of As(III)′s binding mechanisms, showing that, in addition to As(III)−S covalent bonds, noncovalent O⋅⋅⋅As pnictogen bonding plays a vital role. This interaction significantly contributes to the structural stability of the As(III) complexes. Our crystallographic analysis using the PDB database of As(III) S‐adenosylmethionine methyltransferases, augmented by comprehensive theoretical studies including molecular electrostatic potential (MEP), quantum theory of atoms in molecules (QTAIM), and natural bond orbital (NBO) analysis, emphasizes the critical role of pnictogen bonding in these systems. We also undertake a detailed evaluation of the energy characteristics of these pnictogen bonds using various theoretical models. To our knowledge, this is the first time pnictogen bonds in As(III) derivatives have been reported in biological systems, marking a significant advancement in our understanding of arsenic's molecular interactions. [ABSTRACT FROM AUTHOR]

Methanogenic archaea inhabiting anaerobic environments play a crucial role in the global biogeochemical material cycle. The most universal electrogenic reaction of their methane-producing energy metabolism is catalyzed by N 5 -methyl-tetrahydromethanopterin: coenzyme M methyltransferase (MtrABCDEFGH), which couples the vectorial Na+ transport with a methyl transfer between the one-carbon carriers tetrahydromethanopterin and coenzyme M via a vitamin B12 derivative (cobamide) as prosthetic group. We present the 2.08 Å cryo-EM structure of Mtr(ABCDEFG)3 composed of the central Mtr(ABFG)3 stalk symmetrically flanked by three membrane-spanning MtrCDE globes. Tetraether glycolipids visible in the map fill gaps inside the multisubunit complex. Putative coenzyme M and Na+ were identified inside or in a side-pocket of a cytoplasmic cavity formed within MtrCDE. Its bottom marks the gate of the transmembrane pore occluded in the cryo-EM map. By integrating Alphafold2 information, functionally competent MtrA-MtrH and MtrA-MtrCDE subcomplexes could be modeled and thus the methyl-tetrahydromethanopterin demethylation and coenzyme M methylation half-reactions structurally described. Methyl-transfer-driven Na+ transport is proposed to be based on a strong and weak complex between MtrCDE and MtrA carrying vitamin B12, the latter being placed at the entrance of the cytoplasmic MtrCDE cavity. Hypothetically, strongly attached methyl-cob(III)amide (His-on) carrying MtrA induces an inward-facing conformation, Na+ flux into the membrane protein center and finally coenzyme M methylation while the generated loosely attached (or detached) MtrA carrying cob(I)amide (His-off) induces an outward-facing conformation and an extracellular Na+ outflux. Methyl-cob(III)amide (His-on) is regenerated in the distant active site of the methyl-tetrahydromethanopterin binding MtrH implicating a large-scale shuttling movement of the vitamin B12-carrying domain. [ABSTRACT FROM AUTHOR]

The biosyntheses of sodorifien and chlororaphen demonstrate impressively that in bacteria an additional way of terpene biosynthesis was established which results in the production of new – so far unknown – natural terpene products. For these biosynthetic pathways, especially the properties of two specialized enzymes are responsible. This is on the one hand a bifunctional SAM-dependent FPP-methyltransferase with cyclase activity and on the other hand a terpene synthase with altered substrate specificity. Thus, it has been documented that – deviating from the C5 unit-depending isoprene rule – there is another possibility of modifying classical prenyl pyrophosphates to C16 or C17 cyclized substrates. Subsequently, the 136-year-old isoprene dogma has to be expanded or modified. [ABSTRACT FROM AUTHOR]

Flavonoids exhibit various bioactivities including anti-oxidant, anti-tumor, anti-inflammatory, and anti-viral properties. Methylated flavonoids are particularly significant due to their enhanced oral bioavailability, improved intestinal absorption, and greater stability. The heterologous production of plant flavonoids in bacterial factories involves the need for enough biosynthetic precursors to allow for high production levels. These biosynthetic precursors are malonyl-CoA and l-tyrosine. In this work, to enhance flavonoid biosynthesis in Streptomyces albidoflavus, we conducted a transcriptomics study for the identification of candidate genes involved in l-tyrosine catabolism. The hypothesis was that the bacterial metabolic machinery would detect an excess of this amino acid if supplemented with the conventional culture medium and would activate the genes involved in its catabolism towards energy production. Then, by inactivating those overexpressed genes (under an excess of l-tyrosine), it would be possible to increase the intracellular pools of this precursor amino acid and eventually the final flavonoid titers in this bacterial factory. The RNAseq data analysis in the S. albidoflavus wild-type strain highlighted the hppD gene encoding 4-hydroxyphenylpyruvate dioxygenase as a promising target for knock-out, exhibiting a 23.2-fold change (FC) in expression upon l-tyrosine supplementation in comparison to control cultivation conditions. The subsequent knock-out of the hppD gene in S. albidoflavus resulted in a 1.66-fold increase in the naringenin titer, indicating enhanced flavonoid biosynthesis. Leveraging the improved strain of S. albidoflavus, we successfully synthesized the methylated flavanones hesperetin, homoeriodictyol, and homohesperetin, achieving titers of 2.52 mg/L, 1.34 mg/L, and 0.43 mg/L, respectively. In addition, the dimethoxy flavanone homohesperetin was produced as a byproduct of the endogenous metabolism of S. albidoflavus. To our knowledge, this is the first time that hppD deletion was utilized as a strategy to augment the biosynthesis of flavonoids. Furthermore, this is the first report where hesperetin and homoeriodictyol have been synthesized from l-tyrosine as a precursor. Therefore, transcriptomics is, in this case, a successful approach for the identification of catabolism reactions affecting key precursors during flavonoid biosynthesis, allowing the generation of enhanced production strains. [ABSTRACT FROM AUTHOR]

DNA methyltransferase is significant in cellular activities and gene expression, and its aberrant expression is closely linked to various cancers during initiation and progression. Currently, there is a great demand for reliable and label‐free techniques for DNA methyltransferase evaluation in tumor diagnosis and cancer therapy. Herein, a low‐background fluorescent RNA aptamer‐based sensing approach for label‐free quantification of cytosine‐guanine (CpG) dinucleotides methyltransferase (M.SssI) is reported. The fluorogenic light‐up RNA aptamers‐based strategy exhibits high selectivity via restriction endonuclease, padlock‐based recognition, and RNA transcription. By combining rolling circle amplification (RCA), and RNA transcription with fluorescence response of RNA aptamers of Spinach‐dye compound, the proposed platform exhibited efficiently ultrahigh sensitivity toward M.SssI. Eventually, the detection can be achieved in a linear range of 0.02–100 U mL−1 with a detection limit of 1.6 × 10−3 U mL−1. Owing to these superior features, the method is further applied in serum samples spiked M.SssI, which delivers a recovery ranging from 92.0 to 107.0% and a relative standard deviation <7.0%, providing a promising and practical tool for determining M.SssI in complex biological matrices. [ABSTRACT FROM AUTHOR]

Monoterpenoid indole alkaloids (MIA) are one of the largest and most complex alkaloid class in nature, boasting many clinically significant drugs such as anticancer vinblastine and antiarrhythmic ajmaline. Many MIAs undergo nitrogen N-methylation, altering their reactivity and affinity to the biological targets through a straightforward reaction. Remarkably, all known MIA N-methyltransferases (NMT) originate from the neofunctionalization of ancestral γ-tocopherol C-methyltransferases (γTMTs), a phenomenon seemingly unique to the Apocynaceae family. In this study, we unveil and characterize a new γTMT-like enzyme from the plant Tabernaemontana elegans (toad tree): perivine Nβ-methyltransferase (TePeNMT). TePeNMT and other homologs form a distinct clade in our phylogenetic study, setting them apart from other γTMTs and γTMT-like NMTs discovered to date. Enzyme kinetic experiments and enzyme homology modeling studies reveal the significant differences in enzyme active sites between TePeNMT and CrPeNMT, a previously characterized perivine Nβ-methyltransferase from Catharanthus roseus (Madagascar periwinkle). Collectively, our findings suggest that parallel evolution of ancestral γTMTs may be responsible for the occurrence of perivine N-methylation in T. elegans and C. roseus.

Abstract Protein and nucleic acid methylation are important biochemical modifications. In addition to their well-established roles in gene regulation, they also regulate cell signaling, metabolism, and translation. Despite this high biological relevance, little is known about the general regulation of methyltransferase function. Methyltransferases are divided into superfamilies based on structural similarities and further classified into smaller families based on sequence/domain/target similarity. While members within superfamilies differ in substrate specificity, their structurally similar active sites indicate a potential for shared modes of regulation. Growing evidence from one superfamily suggests a common regulatory mode may be through heterooligomerization with other family members. Here, we describe examples of methyltransferase regulation through intrafamily heterooligomerization and discuss how this can be exploited for therapeutic use.

Nsp14 is an RNA methyltransferase (MTase) encoded by all coronaviruses. In fact, many viral families, including DNA viruses, encode MTases that catalyze the methylation of the RNA precap structure, resulting in fully capped viral RNA. This capping is crucial for efficient viral RNA translation, stability, and immune evasion. Our previous research identified nsp14 inhibitors based on the chemical scaffold of its methyl donor − the S-adenosyl methionine (SAM) − featuring a modified adenine base and a substituted arylsulfonamide. However, the binding mode of these inhibitors was based only on docking experiments. To uncover atomic details of nsp14 inhibition we solved the crystal structure of nsp14 bound to STM957. The structure revealed the atomic details of nsp14 inhibition such that the 7-deaza-adenine moiety of STM957 forms specific interactions with Tyr368, Ala353, and Phe367, while the arylsulfonamide moiety engages with Asn388 and Phe506. The large aromatic substituent at the 7-deaza position displaces a network of water molecules near the adenine base. Surprisingly, this was recently observed in the case of an unrelated monkeypox MTase VP39, where the 7-deaza modified SAH analogs also displaced water molecules from the vicinity of the active site.

Exosomes, vesicular structures originating from cells, participate in the conveyance of proteins and nucleic acids. Presently, the centrality of epigenetic modifications in neurological disorders is widely acknowledged. Exosomes exert influence over various epigenetic phenomena, thereby modulating post-transcriptional regulatory processes contingent upon their constituent makeup. Consequently, the heightened attention directed toward exosomes as instigators of epigenetic alterations has burgeoned in recent years. Notably, exosomes serve as vehicles for delivering methyltransferases to recipient cells. More significantly, non-coding RNAs, particularly microRNAs (miRNAs), represent pivotal contents within exosomes, wielding the capacity to influence the expression of diverse factors within the cerebral milieu. The transfer of these exosomal contents amidst brain cells, encompassing neuronal cells and microglia, assumes a critical role in the genesis and progression of neurological disorders, also, this role is not limited to neurological disorders, it may deal with any human disease, such as cancer, and cardiovascular diseases. This review will concentrate on elucidating the regulation of exosome-induced epigenetic events and its subsequent ramifications for neurological diseases. A more profound comprehension of the involvement of exosome-mediated epigenetic regulation in neurological disorders contributes to a heightened awareness of the etiology and advancement of cerebral afflictions. [ABSTRACT FROM AUTHOR]

N6‐methyladenosine (m6A) is an mRNA modification widely found in eukaryotes and plays a crucial role in plant development and stress responses. FIONA1 (FIO1) is a recently identified m6A methyltransferase that regulates Arabidopsis (Arabidopsis thaliana) floral transition; however, its role in stress response remains unknown. In this study, we demonstrate that FIO1‐mediated m6A methylation plays a vital role in salt stress response in Arabidopsis. The loss‐of‐function fio1 mutant was sensitive to salt stress. Importantly, the complementation lines expressing the wild‐type FIO1 exhibited the wild‐type phenotype, whereas the complementation lines expressing the mutant FIO1m, in which two critical amino acid residues essential for methyltransferase activity were mutated, did not recover the wild‐type phenotype under salt stress, indicating that the salt sensitivity is associated with FIO1 methyltransferase activity. Furthermore, FIO1‐mediated m6A methylation regulated ROS production and affected the transcript level of several salt stress‐responsive genes via modulating their mRNA stability in an m6A‐dependent manner in response to salt stress. Importantly, FIO1 is associated with salt stress response by specifically targeting and differentially modulating several salt stress‐responsive genes compared with other m6A writer. Collectively, our findings highlight the molecular mechanism of FIO1‐mediated m6A methylation in the salt stress adaptation. Summary Statement: FIONA1 (FIO1), a recently identified m6A methyltransferase, plays a vital role in salt stress response by modulating the stability of several m6A‐modified salt stress‐responsive genes in Arabidopsis thaliana. [ABSTRACT FROM AUTHOR]

Protein and nucleic acid methylation are important biochemical modifications. In addition to their well-established roles in gene regulation, they also regulate cell signaling, metabolism, and translation. Despite this high biological relevance, little is known about the general regulation of methyltransferase function. Methyltransferases are divided into superfamilies based on structural similarities and further classified into smaller families based on sequence/domain/target similarity. While members within superfamilies differ in substrate specificity, their structurally similar active sites indicate a potential for shared modes of regulation. Growing evidence from one superfamily suggests a common regulatory mode may be through heterooligomerization with other family members. Here, we describe examples of methyltransferase regulation through intrafamily heterooligomerization and discuss how this can be exploited for therapeutic use. [ABSTRACT FROM AUTHOR]

Background: Secondary metabolites such as benzylisoquinoline alkaloids (BIA) have attracted considerable attention because of their pharmacological properties and potential therapeutic applications. Methyltransferases (MTs) can add methyl groups to alkaloid molecules, altering their physicochemical properties and bioactivity, stability, solubility, and recognition by other cellular components. Five types of O-methyltransferases and two types of N-methyltransferases are involved in BIA biosynthesis. Objective: Since MTs may be the source for the discovery and development of novel biomedical, agricultural, and industrial compounds, we performed extensive molecular and phylogenetic analyses of O- and N-methyltransferases in BIA-producing plants. Methods: MTs involved in BIA biosynthesis were isolated from transcriptomes of Berberis koreana and Caulophyllum robustum. We also mined the methyltransferases of Coptis japonica, Papaver somniferum, and Nelumbo nucifera from the National Center for Biotechnology Information protein database. Then, we analyzed the functional motifs and phylogenetic analysis. Result: We mined 42 O-methyltransferases and 8 N-methyltransferases from the five BIA-producing plants. Functional motifs for S-adenosyl-L-methionine-dependent methyltransferases were retained in most methyltransferases, except for the three O-methyltransferases from N. nucifera. Phylogenetic analysis revealed that the methyltransferases were grouped into four clades, I, II, III and IV. The clustering patterns in the phylogenetic analysis suggested a monophyletic origin of methyltransferases and gene duplication within species. The coexistence of different O-methyltransferases in the deep branch subclade might support some cases of substrate promiscuity. Conclusions: Methyltransferases may be a source for the discovery and development of novel biomedical, agricultural, and industrial compounds. Our results contribute to further understanding of their structure and reaction mechanisms, which will require future functional studies. [ABSTRACT FROM AUTHOR]

Deinococcus radiodurans is an extremophilic microorganism that possesses a unique DNA damage repair system, conferring a strong resistance to radiation, desiccation, oxidative stress, and chemical damage. Recently, we discovered that D. radiodurans possesses an N4-methylation (m4C) methyltransferase called M.DraR1, which recognizes the 5′-CCGCGG-3′ sequence and methylates the second cytosine. Here, we revealed its cognate restriction endonuclease R.DraR1 and recognized that it is the only endonuclease specially for non-4C-methylated 5′-CCGCGG-3′ sequence so far. We designated the particular m4C R.DraR1-M.DraR1 as the DraI R-M system. Bioinformatics searches displayed the rarity of the DraI R-M homologous system. Meanwhile, recombination and transformation efficiency experiments demonstrated the important role of the DraI R-M system in response to oxidative stress. In addition, in vitro activity experiments showed that R.DraR1 could exceptionally cleave DNA substrates with a m5C-methlated 5′-CCGCGG-3′ sequence instead of its routine activity, suggesting that this particular R-M component possesses a broader substrate choice. Furthermore, an imbalance of the DraI R-M system led to cell death through regulating genes involved in the maintenance of cell survival such as genome stability, transporter, and energy production. Thus, our research revealed a novel m4C R-M system that plays key roles in maintaining cell viability and defending foreign DNA in D. radiodurans. [ABSTRACT FROM AUTHOR]

Background Clostridium neonatale was isolated during an outbreak of neonatal necrotizing enterocolitis (NEC) in 2002. C. neonatale was validated as a new species within the genus Clostridium sensu stricto in 2018. In the present study, we evaluated the antimicrobial susceptibility, genetic determinants of resistance, and phylogenetic relationships of a collection of clinical isolates of C. neonatale. Methods C. neonatale strains (n = 68) were isolated from the stools of preterm neonates who either developed NEC or were asymptomatic carriers of C. neonatale in different periods and in different hospitals. Antimicrobial susceptibility was determined by the disc diffusion method. The MICs of clindamycin, cefotaxime and tetracycline were determined. Genetic determinants of resistance were screened by PCR (n = 68) and WGS (n = 35). Genotyping of the isolates was performed by MLST. Results Antimicrobial resistance was found to clindamycin (n = 24; 35%), cefotaxime (n = 7; 10%) and tetracycline (n = 1; 1%). One clindamycin-resistant isolate carried erm (B) by PCR. In addition, one isolate carrying tet (M) was tetracycline resistant (MIC = 16 mg/L) and 44 isolates carrying either tet (O), tet (32) or tet (M) were tetracycline susceptible (MICs < 16 mg/L). MLST showed that ST2 and ST15 were significantly associated with tet (32) (P < 0.0001) and tet (O) (P < 0.0001), respectively. From WGS, we identified aph(3′)-IIa and bla TEM-116 genes and a bla CBP-1-like gene. Conclusions C. neonatale is susceptible to anti-anaerobic molecules but resistant to clindamycin, cefotaxime and tetracycline. Genes encoding tetracycline ribosomal protection, macrolide-lincosamide-streptogramin B rRNA methyltransferase, aminoglycoside 3 '- phosphotransferase and β-lactamases have been identified in genomic regions flanked by mobile genetic elements. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) has a high rate of recurrence and poor prognosis, even after curative surgery. Multikinase inhibitors have been applied for HCC patients, but their effect has been restricted. This study aims to clarify the clinical impact of SUV420H1/KMT5B, one of the methyltransferases for histone H4 at lysine 20, and elucidate the novel mechanisms of HCC progression. We retrospectively investigated SUV420H1 expression using HCC clinical tissue samples employing immunohistochemical analysis (n = 350). We then performed loss‐of‐function analysis of SUV420H1 with cell cycle analysis, migration assay, invasion assay and RNA sequence for Gene Ontology (GO) pathway analysis in vitro, and animal experiments with xenograft mice in vivo. The SUV420H1‐high‐score group (n = 154) had significantly poorer prognosis for both 5‐year overall and 2‐year/5‐year disease‐free survival than the SUV420H1‐low‐score group (n = 196) (p < 0.001 and p < 0.05, respectively). The SUV420H1‐high‐score group had pathologically larger tumor size, more tumors, poorer differentiation, and more positive vascular invasion than the SUV420H1‐low‐score group. Multivariate analysis demonstrated that SUV420H1 high score was the poorest independent factor for overall survival. SUV420H1 knockdown could suppress cell cycle from G1 to S phase and cell invasion. GO pathway analysis showed that SUV420H1 contributed to cell proliferation, cell invasion, and/or metastasis. Overexpression of SUV420H1 clinically contributed to poor prognosis in HCC, and the inhibition of SUV420H1 could repress tumor progression and invasion both in vitro and in vivo; thus, further analyses of SUV420H1 are necessary for the discovery of future molecularly targeted drugs. [ABSTRACT FROM AUTHOR]

Background: Arginase 1 (Arg1) encodes a key enzyme that catalyzes the metabolism of arginine to ornithine and urea. In our recent study, we found that knockdown of Arg1 in the lungs of fetal mice induces apoptosis of epithelial cells and dramatically delays initiation of labor. As the most abundant internal mRNA modification, N6‐methyladenosine (m6A) has been found to play important roles in lung development and cellular differentiation. However, if the knockdown of Arg1 affects the RNA m6A modification in fetal lungs remains unknown. Methods: In the current study, the RNA m6A levels and the expression of RNA m6A related enzymes were validated in 13.0 dpc fetal lungs that Arg1 was knocked down by adeno‐associated virus carrying Arg1‐shRNA, using western blot, immunofluorescence, and RT‐qPCR. Results: No statistical differences were found in the expression of methyltransferase, demethylases, and binding proteins in the fetal lungs between AAV‐shArg1‐injected mice and AAV‐2/9‐injected mice. Besides, there is no significant change of overall RNA m6A level in fetal lungs from AAV‐shArg1‐injected mice, compared with that from AAV‐2/9‐injected mice. Conclusions: These results indicate that arginase 1 does not affect RNA m6A methylation in mouse fetal lung, and the mechanisms other than RNA m6A modification underlying the effects of Arg1 knockdown on the fetal lung development and their interaction with labor initiation need to be further explored. [ABSTRACT FROM AUTHOR]

Methylation is one of the critical modifications that regulates numerous biological processes. Guanine capping and methylation at the 7th position (m7G) have been shown to mature mRNA for increased RNA stability and translational efficiency. The m7G capped cap0 RNA remains immature and requires additional methylation at the first nucleotide (N1-2′- O -Me), designated as cap1, to achieve full maturation. This cap1 RNA with N1-2′- O -Me prevents its recognition by innate immune sensors as non-self. Viruses have also evolved various strategies to produce self-like capped RNAs with the N1-2′- O -Me that potentially evades the antiviral response and establishes an efficient replication. In this review, we focus on the importance of the presence of N1-2′- O -Me in viral RNAs and discuss the potential for drug development by targeting host and viral N1-2′- O -methyltransferases. [Display omitted] Tsukamotoet al. summarize the capping mechanisms of viral mRNAs and discuss the potential of drug development targeting viral and host enzymes involved in this process. [ABSTRACT FROM AUTHOR]

Flavonoids are ubiquitous polyphenolic compounds that play a vital role in plants' defense response and medicinal efficacy. UV-B radiation is a vital environmental regulator governing flavonoid biosynthesis in plants. Many plants rapidly biosynthesize flavonoids as a response to UV-B stress conditions. Here, we investigated the effects of flavonoid biosynthesis via UV-B irradiation in Euphorbia lathyris. We found that exposure of the E. lathyris callus to UV-B radiation sharply increased the level of one O-methyltransferase (ElOMT1) transcript and led to the biosynthesis of several methylated flavonoids. The methyltransferase ElOMT1 was expressed heterologously in E. coli, and we tested the catalytic activity of recombinant ElOMT1 with possible substrates, including caffeic acid, baicalin, and luteolin, in vitro. ElOMT1 could efficiently methylate when the hydroxyl groups were contained in the core nucleus of the flavonoid. This molecular characterization identifies a methyltransferase responsible for the chemical modification of the core flavonoid structure through methylation and helps reveal the mechanism of methylated flavonoid biosynthesis in Euphorbiaceae. This study identifies the O-methyltransferase that responds to UV-B irradiation and helps shed light on the mechanism of flavonoid biosynthesis in Euphorbia lathyris. [ABSTRACT FROM AUTHOR]

Objective To investigate the effects of overexpression and knockdown of methyltransferase-like protein 14 (METTL14) on proliferation, apoptosis, invasion and migration of acute promyelocytic leukemia cells. Methods Human acute promyelocytic leukemia cells NB4 were cultured, and were divided into the overexpression control (OverNC) group, overexpression group, interference control (si-NC) group, and interference group, respectively. The blank plasmid and METTL14 plasmid were transfected into NB4 cells in the control group and the overexpression group, respectively. The si-NC and si-METTL14 were transfected into NB4 cells in the interference control group and the interference group, respectively. Cell proliferation was detected by CCK-8, apoptosis was detected by flow cytometry, cell invasion and migration abilities were detected by Transwell chamber, and N-6 methyladenine (m6A) modification level was detected by Dot blotting assay. Results Compared with the Over-NC group, the cell proliferation rate increased, apoptosis rate decreased, cell migration number and cell invasion number increased, and m6A modification level of NB4 cells increased in the overexpression group (all P<0.05). Compared with the si-NC group, the proliferation rate decreased, apoptosis rate increased, cell migration number and cell invasion number decreased, and the m6A modification level of NB4 cells decreased in the interference group (all P<0.05). Conclusion METTL14 can promote the proliferation, migration and invasion of NB4 cells, and its mechanism may be related to the increase of m6A modification level. [ABSTRACT FROM AUTHOR]

Autophagy is an important mechanism to maintain cellular function and metabolism, whereas abnormal autophagy can cause the advent and worsening of various diseases. N6-Methyladenosine（ m6A） RNA methylation is a reversible RNA modification, which is regulated by m6A methyltransferase, m6A demethylase and m6A-binding protein. Studies have shown that autophagy-related genes promote or attenuate autophagy level dependent on the regulation of m6A, and then participate in the process of diseases. This paper reviews the progress of m6A modification regulatory enzymes and their binding proteins in regulating cell autophagy to provide reference for future researches. [ABSTRACT FROM AUTHOR]

IntroductionThe African oil palm (Elaeis guineensis Jacq.) is the predominant oil crop in the world. In addition to triacylglycerols, crude palm oil (CPO) extracted from the mesocarp of the fruits, contains high amounts of provitamin A (carotenes) and vitamin E (tocochromanols). Because of their unsaturated nature, the carotenes are prone to oxidation and therefore are in part limiting for the shelf life of CPO.MethodsA tree with unusual toochromanol composition was identified by HPLC screening of the mesocarp of wild trees. Polymorphisms in a candidate gene were identified by DNA sequencing. The candidate protein was heterologously expressed in Escherichia coli coli and Arabidopsis thaliana to test for enzyme activity. Oxidative stability of the CPO was studied by following carotene degradation over time.ResultsIn the present study, a wild Oil Palm tree (C59) from Cameroon was identified that lacks α-tocopherol and α-tocotrienol and instead accumulates the respective γ forms, suggesting that the activity of γ-tocopherol methyltransferase (VTE4) was affected. Sequencing of the VTE4 locus in the genome of plant C59 identified a G/C polymorphism that causes the exchange of a highly conserved tryptophan at position 290 with serine. The W290S exchange renders the VTE4 enzyme inactive, as shown after expression in Escherichia coli and Arabidopsis thaliana. The oxidative stability of carotenes in the mesocarp of the wild palm C59 was enhanced compared with control accessions. Furthermore, supplementation of commercial palm oil with different tocochromanols showed that γ-tocotrienol exerts a stronger effect during the protection of carotenes against oxidation than α-tocotrienol.DiscussionTherefore, the introduction of the high γ-tocotrienol trait into elite breeding lines represents a potent strategy to protect carotenes against oxidation and extend the shelf life of CPO, hence allowing the development of a value added high-carotene CPO to be used to fight against vitamin A deficiency.