Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 20 de 8.245
Filter
1.
Methods Enzymol ; 680: 101-137, 2023.
Article in English | MEDLINE | ID: mdl-36710008

ABSTRACT

Phospholipids play an essential role as a barrier between cell content and the extracellular environment and regulate various cell signaling processes. Phosphatidylcholine (PtdCho) is one of the most abundant phospholipids in plant, animal, and some prokaryote cell membranes. In plants and some parasites, the biosynthesis of PtdCho begins with the amino acid serine, followed mainly through a phosphoethanolamine N-methyltransferase (PMT)-mediated biosynthetic pathway to phosphocholine (pCho). Because the PMT-mediated pathway, referred to as the phosphobase methylation pathway, produces a series of important primary and specialized metabolites for plant development and stress response, understanding the PMT enzyme is a key aspect of engineering plants with improved stress tolerance and fortified nutrients. Importantly, given the very limited phylogenetic distribution of PMTs, functional analysis and the identification of inhibitors targeting PMTs have potential and positive impacts in humans and in veterinary and agricultural fields. Here, we describe detailed basic knowledge and practical research methods to enable the systematic study of the biochemical and biophysical functions of PMT. The research methods described in this chapter are also applicable to the studies of other ubiquitous S-adenosyl-l-methionine (SAM)-dependent methyltransferases in all kingdoms.


Subject(s)
Methyltransferases , Parasites , Humans , Animals , Methyltransferases/metabolism , S-Adenosylmethionine/metabolism , Choline , Parasites/metabolism , Phylogeny , Phospholipids
2.
Zhonghua Er Bi Yan Hou Tou Jing Wai Ke Za Zhi ; 57(12): 1470-1478, 2022 Dec 07.
Article in Chinese | MEDLINE | ID: mdl-36707952

ABSTRACT

Objective: To investigate the roles of N6-methyladenosine (m6A) modification in regulating RP11-426A6.5 in the development of laryngeal squamous cell carcinoma (LSCC). Methods: The methylation and expression levels of lncRNAs were identified and important lncRNAs were screened utilizing long non-coding RNA (lncRNA) m6A methylation microarray. Cancer and para cancer tissue samples were taken from 48 LSCC patients hospitalized to the Department of Otolaryngology-Head and Neck Surgery of the Second Affiliated Hospital of Harbin Medical University between January and September 2017. Expression profiling microarray was performed in 3 of 48 LSCC samples, and methylated RNA immunoprecipitation-quantitative PCR (MeRIP-qPCR) and quantitative real-time fluorescent PCR (qRT-PCR) were performed in the remaining 45 LSCC samples to verify the m6A modification and expression levels of RP11-426A6.5. Correlations between RP11-426A6.5 and clinical factors were anlysed. Laryngeal cancer cell line with low expression of RP11-426A6.5 was created in vitro using RNA interference (RNAi) technology. The 5-Ethynyl-2'-deoxyuridine (EdU) cell proliferation experiment, wound healing experiment, and transwell invasion experiment were used respectively to measure the proliferation, migration, and invasion of LSCC cells. The effect of RP11-426A6.5 down-regulation on the growth of transplanted tumors in vivo was verified by nude mice tumorigenesis assay. The Cancer Genome Atlas (TCGA) database and sequence-based RNA adenosine methylation site predictor (SRAMP) website were used to predict the enzymes and corresponding methylation sites. MazF digestion was chosen to validate the binding sites. RNAi technology was used to observe the changes in cell function after interfering with the expression of the corresponding genes of the modified enzymes. MeRIP-qPCR was used to detect the level of RP11-426A6.5 m6A cell line treated with actinomycin D was used to observe the stability of RP11-426A6.5. Results: RP11-426A6.5 methylation and expression levels were significantly higher in LSCC tissues than those in paracancerous tissues (methylation levels: 23.828±4.975 vs 20.280±3.607; expression levels: 1.197±0.314 vs 1.015±0.170, all P values<0.05). RP11-426A6.5 expression levels were closely correlated with T stage (T1-2: 1.081±0.298 vs T3-4: 1.306±0.292, χ2=5.35, P<0.05). The postoperative survival of patients with high RP11-426A6.5 expressions was significantly lower than that of patients with low RP11-426A6.5 expression (P=0.046). Assays in vitro and in vivo showed that the downregulation of RP11-426A6.5 significantly decreased the proliferation, migration, and invasion abilities of LSCC cells and the growth of transplanted tumors. The binding of methyltransferase-like 3 (METTL3), an m6A-modified enzyme, to the corresponding methylation site of RP11-426A6.5 enhanced its stability and mediated its regulation of malignant behaviors of LSCC cells. Conclusions: RP11-426A6.5 can regulate the malignant behaviors of LSCC cells, which is mediated by the m6A modification process involving in the methyltransferase METTL3.


Subject(s)
Head and Neck Neoplasms , Laryngeal Neoplasms , MicroRNAs , RNA, Long Noncoding , Animals , Mice , Squamous Cell Carcinoma of Head and Neck/genetics , RNA, Long Noncoding/genetics , Mice, Nude , Laryngeal Neoplasms/pathology , Cell Proliferation/genetics , Methyltransferases/genetics , Methyltransferases/metabolism , Gene Expression Regulation, Neoplastic , Cell Line, Tumor , MicroRNAs/genetics
3.
Autoimmunity ; 56(1): 2167983, 2023 Dec.
Article in English | MEDLINE | ID: mdl-36708146

ABSTRACT

Skin diseases are global health issues caused by multiple pathogenic factors, in which epigenetics plays an invaluable role. Post-transcriptional RNA modifications are important epigenetic mechanism that regulate gene expression at the genome-wide level. N6-methyladenosine (m6A) is the most prevalent modification that occurs in the messenger RNAs (mRNA) of most eukaryotes, which is installed by methyltransferases called "writers", removed by demethylases called "erasers", and recognised by RNA-binding proteins called "readers". To date, m6A is emerging to play essential part in both physiological processes and pathological progression, including skin diseases. However, a systematic summary of m6A in skin disease has not yet been reported. This review starts by illustrating each m6A-related modifier specifically and their roles in RNA processing, and then focus on the existing research advances of m6A in immune homeostasis and skin diseases.


Subject(s)
Methyltransferases , Skin Diseases , Humans , Methylation , Methyltransferases/genetics , Methyltransferases/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Adenosine/genetics , Adenosine/metabolism , Skin Diseases/genetics , RNA/metabolism
4.
Proc Natl Acad Sci U S A ; 120(5): e2214684120, 2023 Jan 31.
Article in English | MEDLINE | ID: mdl-36693099

ABSTRACT

Embryo implantation, a crucial step in human reproduction, is tightly controlled by estrogen and progesterone (P4) via estrogen receptor alpha and progesterone receptor (PGR), respectively. Here, we report that N6-methyladenosine (m6A), the most abundant mRNA modification in eukaryotes, plays an essential role in embryo implantation through the maintenance of P4 signaling. Conditional deletion of methyltransferase-like 3 (Mettl3), encoding the m6A writer METTL3, in the female reproductive tract using a Cre mouse line with Pgr promoter (Pgr-Cre) resulted in complete implantation failure due to pre-implantation embryo loss and defective uterine receptivity. Moreover, the uterus of Mettl3 null mice failed to respond to artificial decidualization. We further found that Mettl3 deletion was accompanied by a marked decrease in PGR protein expression. Mechanistically, we found that Pgr mRNA is a direct target for METTL3-mediated m6A modification. A luciferase assay revealed that the m6A modification in the 5' untranslated region (5'-UTR) of Pgr mRNA enhances PGR protein translation efficiency in a YTHDF1-dependent manner. Finally, we demonstrated that METTL3 is required for human endometrial stromal cell decidualization in vitro and that the METTL3-PGR axis is conserved between mice and humans. In summary, this study provides evidence that METTL3 is essential for normal P4 signaling during embryo implantation via m6A-mediated translation control of Pgr mRNA.


Subject(s)
Progesterone , Receptors, Progesterone , Female , Mice , Humans , Animals , Progesterone/metabolism , Receptors, Progesterone/genetics , Receptors, Progesterone/metabolism , Embryo Implantation/genetics , Uterus/metabolism , Methyltransferases/genetics , Methyltransferases/metabolism , Mice, Knockout , RNA, Messenger/metabolism
5.
Viruses ; 15(1)2023 Jan 10.
Article in English | MEDLINE | ID: mdl-36680231

ABSTRACT

Since late 2016, a yellow fever virus (YFV) variant carrying a set of nine amino acid variations has circulated in South America. Three of them were mapped on the methyltransferase (MTase) domain of viral NS5 protein. To assess whether these changes affected viral infectivity, we synthesized YFV carrying the MTase of circulating lineage as well as its isoform with the residues of the previous strains (NS5 K101R, NS5 V138I, and NS5 G173S). We observed a slight difference in viral growth properties and plaque phenotype between the two synthetic YFVs. However, the MTase polymorphisms associated with the Brazilian strain of YFV (2016-2019) confer more susceptibility to the IFN-I. In addition, in vitro MTase assay revealed that the interaction between the YFV MTase and the methyl donor molecule (SAM) is altered in the Brazilian MTase variant. Altogether, the results reported here describe that the MTase carrying the molecular signature of the Brazilian YFV circulating since 2016 might display a slight decrease in its catalytic activity but virtually no effect on viral fitness in the parameters comprised in this study. The most marked influence of these residues stands in the immune escape against the antiviral response mediated by IFN-I.


Subject(s)
Interferon Type I , Yellow fever virus , Yellow fever virus/physiology , Interferon Type I/genetics , Amino Acids , Immune Evasion , Brazil , Methyltransferases/metabolism , Viral Nonstructural Proteins/genetics
6.
Int J Mol Sci ; 24(2)2023 Jan 10.
Article in English | MEDLINE | ID: mdl-36674842

ABSTRACT

The PRDM family of methyltransferases has been implicated in cellular proliferation and differentiation and is deregulated in human diseases, most notably in cancer. PRDMs are related to the SET domain family of methyltransferases; however, from the 19 PRDMs only a few PRDMs with defined enzymatic activities are known. PRDM15 is an uncharacterized transcriptional regulator, with significant structural disorder and lack of defined small-molecule binding pockets. Many aspects of PRDM15 are yet unknown, including its structure, substrates, reaction mechanism, and its methylation profile. Here, we employ a series of computational approaches for an exploratory investigation of its potential substrates and reaction mechanism. Using the knowledge of PRDM9 and current knowledge of PRDM15 as basis, we tried to identify genuine substrates of PRDM15. We start from histone-based peptides and learn that the native substrates of PRDM15 may be non-histone proteins. In the future, a combination of sequence-based approaches and signature motif analysis may provide new leads. In summary, our results provide new information about the uncharacterized methyltransferase, PRDM15.


Subject(s)
Methyltransferases , Neoplasms , Humans , Methyltransferases/metabolism , Methylation , Histones/genetics , Histones/metabolism , DNA-Binding Proteins/metabolism , Transcription Factors/genetics , Transcription Factors/metabolism , Histone-Lysine N-Methyltransferase/metabolism
7.
Int J Mol Sci ; 24(2)2023 Jan 10.
Article in English | MEDLINE | ID: mdl-36674903

ABSTRACT

Methylation of histone 3 at lysine 79 (H3K79) and its catalyst, a disrupter of telomeric silencing (DOT1l), have been coupled to multiple forms of stress, such as bioenergetic and ER challenges. However, studies on H3K79 methylation and Dot1l in the (aging) brain and neurons are limited. This, together with the increasing evidence of a dynamic neuroepigenome, made us wonder if H3K79 methylation and its activator Dot1l could play important roles in brain aging and associated disorders. In aged humans, we found strong and consistent global hypermethylation of H3K79 in neurons. Specific in dopaminergic neurons, we found a strong increase in H3K79 methylation in lipofucsin positive neurons, which are linked to pathology. In animals, where we conditionally removed Dot1l, we found a rapid loss of H3K79 methylation. As a consequence, we found some decrease in specific dopaminergic genes, and surprisingly, a clear up-regulation of almost all genes belonging to the family of the respiratory chain. These data, in relation to the observed increase in global H3K79 methylation, suggest that there is an inverse relationship between H3K79 methylation and the capacity of energy metabolism in neuronal systems.


Subject(s)
Genes, Mitochondrial , Methyltransferases , Animals , Humans , Aged , Methyltransferases/metabolism , Histones/genetics , Histones/metabolism , DNA Methylation , Neurons/metabolism , Brain/metabolism , Histone-Lysine N-Methyltransferase/genetics , Histone-Lysine N-Methyltransferase/metabolism
8.
Int J Mol Sci ; 24(2)2023 Jan 11.
Article in English | MEDLINE | ID: mdl-36674918

ABSTRACT

Excessive differentiation of osteoclasts contributes to the disruption of bone homeostasis in inflammatory bone diseases. Methyltransferase-like 3 (METTL3), the core methyltransferase that installs an N6-methyladenosine (m6A) modification on RNA, has been reported to participate in bone pathophysiology. However, whether METTL3-mediated m6A affects osteoclast differentiation in inflammatory conditions remains unelucidated. In this study, we observed that the total m6A content and METTL3 expression decreased during LPS-induced osteoclastogenesis. After knocking down METTL3, we found reduced levels of the number of osteoclasts, osteoclast-related gene expression and bone resorption area. A METTL3 deficiency increased osteoclast apoptosis and pro-apoptotic protein expression. RNA sequencing analysis showed that differentially expressed genes in METTL3-deficient cells were mainly associated with the mitochondrial function. The expression of the mitochondrial function-related genes, ATP production and mitochondrial membrane potential decreased after METTL3 knockdown. Moreover, the most obviously upregulated gene in RNA-Seq was Nos2, which encoded the iNOS protein to induce nitric oxide (NO) synthesis. METTL3 knockdown increased the levels of Nos2 mRNA, iNOS protein and NO content. NOS inhibitor L-NAME rescued the inhibited mitochondrial function and osteoclast formation while suppressing osteoclast apoptosis in METTL3-silenced cells. Mechanistically, a METTL3 deficiency promoted the stability and expression of Nos2 mRNA, and similar results were observed after m6A-binding protein YTHDF1 knockdown. Further in vivo evidence revealed that METTL3 knockdown attenuated the inflammatory osteolysis of the murine calvaria and suppressed osteoclast formation. In conclusion, these data suggested that METTL3 knockdown exacerbated iNOS/NO-mediated mitochondrial dysfunction by promoting a Nos2 mRNA stability in a YTHDF1-dependent manner and further inhibited osteoclast differentiation and increased osteoclast apoptosis in inflammatory conditions.


Subject(s)
Bone Resorption , Osteoclasts , Mice , Animals , Osteoclasts/metabolism , Nitric Oxide/metabolism , Bone Resorption/metabolism , Methyltransferases/genetics , Methyltransferases/metabolism , RNA, Messenger/genetics
9.
Proc Natl Acad Sci U S A ; 120(4): e2208941120, 2023 Jan 24.
Article in English | MEDLINE | ID: mdl-36656859

ABSTRACT

p97 is an essential AAA+ ATPase that extracts and unfolds substrate proteins from membranes and protein complexes. Through its mode of action, p97 contributes to various cellular processes, such as membrane fusion, ER-associated protein degradation, DNA repair, and many others. Diverse p97 functions and protein interactions are regulated by a large number of adaptor proteins. Alveolar soft part sarcoma locus (ASPL) is a unique adaptor protein that regulates p97 by disassembling functional p97 hexamers to smaller entities. An alternative mechanism to regulate the activity and interactions of p97 is by posttranslational modifications (PTMs). Although more than 140 PTMs have been identified in p97, only a handful of those have been described in detail. Here we present structural and biochemical data to explain how the p97-remodeling adaptor protein ASPL enables the metastasis promoting methyltransferase METTL21D to bind and trimethylate p97 at a single lysine side chain, which is deeply buried inside functional p97 hexamers. The crystal structure of a heterotrimeric p97:ASPL:METTL21D complex in the presence of cofactors ATP and S-adenosyl homocysteine reveals how structural remodeling by ASPL exposes the crucial lysine residue of p97 to facilitate its trimethylation by METTL21D. The structure also uncovers a role of the second region of homology (SRH) present in the first ATPase domain of p97 in binding of a modifying enzyme to the AAA+ ATPase. Investigation of this interaction in the human, fish, and plant reveals fine details on the mechanism and significance of p97 trimethylation by METTL21D across different organisms.


Subject(s)
ATPases Associated with Diverse Cellular Activities , Adenosine Triphosphatases , Methyltransferases , Animals , Humans , Adaptor Proteins, Signal Transducing/metabolism , Adenosine Triphosphatases/metabolism , ATPases Associated with Diverse Cellular Activities/metabolism , Lysine/metabolism , Methylation , Protein Binding , Protein Processing, Post-Translational , Transcription Factors/metabolism , Valosin Containing Protein/metabolism , Methyltransferases/metabolism
10.
J Cell Sci ; 136(2)2023 Jan 15.
Article in English | MEDLINE | ID: mdl-36647772

ABSTRACT

N-terminal methylation of the α-amine group (Nα-methylation) is a post-translational modification (PTM) that was discovered over 40 years ago. Although it is not the most abundant of the Nα-PTMs, there are more than 300 predicted substrates of the three known mammalian Nα-methyltransferases, METTL11A and METTL11B (also known as NTMT1 and NTMT2, respectively) and METTL13. Of these ∼300 targets, the bulk are acted upon by METTL11A. Only one substrate is known to be Nα-methylated by METTL13, and METTL11B has no proven in vivo targets or predicted targets that are not also methylated by METTL11A. Given that METTL11A could clearly handle the entire substrate burden of Nα-methylation, it is unclear why three distinct Nα-methyltransferases have evolved. However, recent evidence suggests that many methyltransferases perform important biological functions outside of their catalytic activity, and the Nα-methyltransferases might be part of this emerging group. Here, we describe the distinct expression, localization and physiological roles of each Nα-methyltransferase, and compare these characteristics to other methyltransferases with non-catalytic functions, as well as to methyltransferases with both catalytic and non-catalytic functions, to give a better understanding of the global roles of these proteins. Based on these comparisons, we hypothesize that these three enzymes do not just have one common function but are actually performing three unique jobs in the cell.


Subject(s)
Mammals , Methyltransferases , Animals , Methyltransferases/genetics , Methyltransferases/metabolism , Methylation , Catalysis , Mammals/metabolism
11.
Commun Biol ; 6(1): 54, 2023 Jan 16.
Article in English | MEDLINE | ID: mdl-36646841

ABSTRACT

The 22nd genetically encoded amino acid, pyrrolysine, plays a unique role in the key step in the growth of methanogens on mono-, di-, and tri-methylamines by activating the methyl group of these substrates for transfer to a corrinoid cofactor. Previous crystal structures of the Methanosarcina barkeri monomethylamine methyltransferase elucidated the structure of pyrrolysine and provide insight into its role in monomethylamine activation. Herein, we report the second structure of a pyrrolysine-containing protein, the M. barkeri trimethylamine methyltransferase MttB, and its structure bound to sulfite, a substrate analog of trimethylamine. We also report the structure of MttB in complex with its cognate corrinoid protein MttC, which specifically receives the methyl group from the pyrrolysine-activated trimethylamine substrate during methanogenesis. Together these structures provide key insights into the role of pyrrolysine in methyl group transfer from trimethylamine to the corrinoid cofactor in MttC.


Subject(s)
Corrinoids , Methyltransferases , Methyltransferases/metabolism , Methylamines/metabolism , Corrinoids/metabolism
12.
Int J Biol Sci ; 19(2): 705-720, 2023.
Article in English | MEDLINE | ID: mdl-36632456

ABSTRACT

Autophagy is an evolutionarily conserved cellular degradation and recycling process. It is important for maintaining vital cellular function and metabolism. Abnormal autophagy activity can cause the development of various diseases. N6-methyladenosine (m6A) methylation is the most prevalent and abundant internal modification in eukaryotes, affecting almost all aspects of RNA metabolism. The process of m6A modification is dynamic and adjustable. Its regulation depends on the regulation of m6A methyltransferases, m6A demethylases, and m6A binding proteins. m6A methylation and autophagy are two crucial and independent cellular events. Recent studies have shown that m6A modification mediates the transcriptional and post-transcriptional regulation of autophagy-related genes, affecting autophagy regulatory networks in multiple diseases. However, the regulatory effects of m6A regulators on autophagy in human diseases are not adequately acknowledged. In the present review, we summarized the latest knowledge of m6A modification in autophagy and elucidated the molecular regulatory mechanisms underlying m6A modification in autophagy regulatory networks. Moreover, we discuss the potentiality of m6A regulators serving as promising predictive biomarkers for human disease diagnosis and targets for therapy. This review will increase our understanding of the relationship between m6A methylation and autophagy, and provide novel insights to specifically target m6A modification in autophagy-associated therapeutic strategies.


Subject(s)
Gene Expression Regulation , Methyltransferases , Humans , Methylation , Methyltransferases/genetics , Methyltransferases/metabolism , Autophagy/genetics
13.
Int J Mol Sci ; 24(1)2022 Dec 22.
Article in English | MEDLINE | ID: mdl-36613611

ABSTRACT

Haploinsufficiency of the SETD5 gene, encoding a SET domain-containing histone methyltransferase, has been identified as a cause of intellectual disability and Autism Spectrum Disorder (ASD). Recently, the zebrafish has emerged as a valuable model to study neurodevelopmental disorders because of its genetic tractability, robust behavioral traits and amenability to high-throughput drug screening. To model human SETD5 haploinsufficiency, we generated zebrafish setd5 mutants using the CRISPR/Cas9 technology and characterized their morphological, behavioral and molecular phenotypes. According to our observation that setd5 is expressed in adult zebrafish brain, including those areas controlling social behavior, we found that setd5 heterozygous mutants exhibit defective aggregation and coordination abilities required for shoaling interactions, as well as indifference to social stimuli. Interestingly, impairment in social interest is rescued by risperidone, an antipsychotic drug used to treat behavioral traits in ASD individuals. The molecular analysis underscored the downregulation of genes encoding proteins involved in the synaptic structure and function in the adult brain, thus suggesting that brain hypo-connectivity could be responsible for the social impairments of setd5 mutant fishes. The zebrafish setd5 mutants display ASD-like features and are a promising setd5 haploinsufficiency model for drug screening aimed at reversing the behavioral phenotypes.


Subject(s)
Autism Spectrum Disorder , Methyltransferases , Animals , Humans , Autism Spectrum Disorder/genetics , Autism Spectrum Disorder/metabolism , Brain/metabolism , CRISPR-Cas Systems , Methyltransferases/genetics , Methyltransferases/metabolism , Zebrafish/genetics , Zebrafish/metabolism , Zebrafish Proteins/genetics , Zebrafish Proteins/metabolism , Social Behavior
14.
Int J Mol Sci ; 24(1)2023 Jan 01.
Article in English | MEDLINE | ID: mdl-36614187

ABSTRACT

Ladderane lipids (found in the membranes of anaerobic ammonium-oxidizing [anammox] bacteria) have unique ladder-like hydrophobic groups, and their highly strained exotic structure has attracted the attention of scientists. Although enzymes encoded in type II fatty acid biosynthesis (FASII) gene clusters in anammox bacteria, such as S-adenosyl-l-methionine (SAM)-dependent enzymes, have been proposed to construct a ladder-like structure using a substrate connected to acyl carrier protein from anammox bacteria (AmxACP), no experimental evidence to support this hypothesis was reported to date. Here, we report the crystal structure of a SAM-dependent methyltransferase from anammox bacteria (AmxMT1) that has a substrate and active site pocket between a class I SAM methyltransferase-like core domain and an additional α-helix inserted into the core domain. Structural comparisons with homologous SAM-dependent C-methyltransferases in polyketide synthase, AmxACP pull-down assays, AmxACP/AmxMT1 complex structure predictions by AlphaFold, and a substrate docking simulation suggested that a small compound connected to AmxACP could be inserted into the pocket of AmxMT1, and then the enzyme transfers a methyl group from SAM to the substrate to produce branched lipids. Although the enzymes responsible for constructing the ladder-like structure remain unknown, our study, for the first time, supports the hypothesis that biosynthetic intermediates connected to AmxACP are processed by SAM-dependent enzymes, which are not typically involved in the FASII system, to produce the ladder-like structure of ladderane lipids in anammox bacteria.


Subject(s)
Methionine , S-Adenosylmethionine , S-Adenosylmethionine/metabolism , Methionine/metabolism , Acyl Carrier Protein/metabolism , Methyltransferases/metabolism , Anaerobic Ammonia Oxidation , Bacteria/metabolism , Racemethionine/metabolism , Lipids , Bacterial Proteins/genetics , Bacterial Proteins/metabolism
15.
Int J Mol Sci ; 24(1)2023 Jan 01.
Article in English | MEDLINE | ID: mdl-36614216

ABSTRACT

N6-metyladenosine (m6A), one of the most common RNA methylation modifications in mammals, has attracted extensive attentions owing to its regulatory roles in a variety of physiological and pathological processes. As a reversible epigenetic modification on RNAs, m6A is dynamically mediated by the functional interplay among the regulatory proteins of methyltransferases, demethylases and methyl-binding proteins. In recent years, it has become increasingly clear that m6A modification is associated with the production and function of microRNAs (miRNAs). In this review, we summarize the specific kinds of m6A modification methyltransferases, demethylases and methyl-binding proteins. In particular, we focus on describing the roles of m6A modification and its regulatory proteins in the production and function of miRNAs in a variety of pathological and physiological processes. More importantly, we further discuss the mediating mechanisms of miRNAs in m6A modification and its regulatory proteins during the occurrence and development of various diseases.


Subject(s)
MicroRNAs , Animals , MicroRNAs/genetics , MicroRNAs/metabolism , Adenosine/metabolism , Methylation , Methyltransferases/metabolism , Epigenesis, Genetic , Carrier Proteins/metabolism , Transcription Factors/metabolism , Mammals/metabolism
16.
Molecules ; 28(1)2022 Dec 21.
Article in English | MEDLINE | ID: mdl-36615239

ABSTRACT

In plants, methylation is a common step in specialized metabolic pathways, leading to a vast diversity of natural products. The methylation of these small molecules is catalyzed by S-adenosyl-l-methionine (SAM)-dependent methyltransferases, which are categorized based on the methyl-accepting atom (O, N, C, S, or Se). These methyltransferases are responsible for the transformation of metabolites involved in plant defense response, pigments, and cell signaling. Plant natural product methyltransferases are part of the Class I methyltransferase-superfamily containing the canonical Rossmann fold. Recent advances in genomics have accelerated the functional characterization of plant natural product methyltransferases, allowing for the determination of substrate specificities and regioselectivity and further realizing the potential for enzyme engineering. This review compiles known biochemically characterized plant natural product methyltransferases that have contributed to our knowledge in the diversification of small molecules mediated by methylation steps.


Subject(s)
Biological Products , Methyltransferases , Methyltransferases/metabolism , Methylation , Plants/genetics , Plants/metabolism , S-Adenosylmethionine/metabolism , Substrate Specificity
17.
Cell Stem Cell ; 30(1): 52-68.e13, 2023 Jan 05.
Article in English | MEDLINE | ID: mdl-36608679

ABSTRACT

N6-methyladenosine (m6A), the most prevalent internal modification in mammalian mRNAs, is involved in many pathological processes. METTL16 is a recently identified m6A methyltransferase. However, its role in leukemia has yet to be investigated. Here, we show that METTL16 is a highly essential gene for the survival of acute myeloid leukemia (AML) cells via CRISPR-Cas9 screening and experimental validation. METTL16 is aberrantly overexpressed in human AML cells, especially in leukemia stem cells (LSCs) and leukemia-initiating cells (LICs). Genetic depletion of METTL16 dramatically suppresses AML initiation/development and maintenance and significantly attenuates LSC/LIC self-renewal, while moderately influencing normal hematopoiesis in mice. Mechanistically, METTL16 exerts its oncogenic role by promoting expression of branched-chain amino acid (BCAA) transaminase 1 (BCAT1) and BCAT2 in an m6A-dependent manner and reprogramming BCAA metabolism in AML. Collectively, our results characterize the METTL16/m6A/BCAT1-2/BCAA axis in leukemogenesis and highlight the essential role of METTL16-mediated m6A epitranscriptome and BCAA metabolism reprograming in leukemogenesis and LSC/LIC maintenance.


Subject(s)
Cell Self Renewal , Leukemia, Myeloid, Acute , Mice , Humans , Animals , Leukemia, Myeloid, Acute/pathology , Carcinogenesis/pathology , RNA, Messenger/metabolism , Amino Acids, Branched-Chain/genetics , Amino Acids, Branched-Chain/metabolism , Neoplastic Stem Cells/pathology , Mammals/metabolism , Transaminases/genetics , Transaminases/metabolism , Methyltransferases/genetics , Methyltransferases/metabolism
18.
J Exp Clin Cancer Res ; 42(1): 10, 2023 Jan 07.
Article in English | MEDLINE | ID: mdl-36609396

ABSTRACT

BACKGROUND: Posttranscriptional modification of tumor-associated factors plays a pivotal role in breast cancer progression. However, the underlying mechanism remains unknown. M6A modifications in cancer cells are dynamic and reversible and have been found to impact tumor initiation and progression through various mechanisms. In this study, we explored the regulatory mechanism of breast cancer cell proliferation and metabolism through m6A methylation in the Hippo pathway.  METHODS: A combination of MeRIP-seq, RNA-seq and metabolomics-seq was utilized to reveal a map of m6A modifications in breast cancer tissues and cells. We conducted RNA pull-down assays, RIP-qPCR, MeRIP-qPCR, and RNA stability analysis to identify the relationship between m6A proteins and LATS1 in m6A regulation in breast cancer cells. The expression and biological functions of m6A proteins were confirmed in breast cancer cells in vitro and in vivo. Furthermore, we investigated the phosphorylation levels and localization of YAP/TAZ to reveal that the activity of the Hippo pathway was affected by m6A regulation of LATS1 in breast cancer cells.  RESULTS: We demonstrated that m6A regulation plays an important role in proliferation and glycolytic metabolism in breast cancer through the Hippo pathway factor, LATS1. METTL3 was identified as the m6A writer, with YTHDF2 as the reader protein of LATS1 mRNA, which plays a positive role in promoting both tumorigenesis and glycolysis in breast cancer. High levels of m6A modification were induced by METTL3 in LATS1 mRNA. YTHDF2 identified m6A sites in LATS1 mRNA and reduced its stability. Knockout of the protein expression of METTL3 or YTHDF2 increased the expression of LATS1 mRNA and suppressed breast cancer tumorigenesis by activating YAP/TAZ in the Hippo pathway. CONCLUSIONS: In summary, we discovered that the METTL3-LATS1-YTHDF2 pathway plays an important role in the progression of breast cancer by activating YAP/TAZ in the Hippo pathway.


Subject(s)
Breast Neoplasms , Humans , Female , Methylation , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Cell Transformation, Neoplastic/genetics , Carcinogenesis/genetics , Transcription Factors/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Methyltransferases/genetics , Methyltransferases/metabolism
19.
Life Sci ; 315: 121359, 2023 Feb 15.
Article in English | MEDLINE | ID: mdl-36608868

ABSTRACT

AIMS: Previous studies have shown that RNA binding motif 10 (RBM10) is a potential tumor suppressor protein that can inhibit proliferation and promote apoptosis of non-small cell lung cancer (NSCLC). Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) plays an important role in promoting the development of lung cancer. Inhibiting its m6A methylation can effectively inhibit the invasion and metastasis of lung cancer. There is concern that RBM10 could affect MALAT1 m6A methylation for the invasion and migration of NSCLC. MAIN METHODS AND FINDINGS: Transwell and wound healing assays showed that RBM10 significantly inhibited the invasion and migration of NSCLC. CLIP-Seq showed that among all RBM10 binding RNAs, MALAT1 had the highest binding peak among all non-coding RNAs. RNA immunoprecipitation verified the direct combination of RBM10 and MALAT1. The rescue experiment confirmed that RBM10 affected the phosphorylation of the PI3K/AKT/mTOR pathway protein as well as the invasion and migration ability by regulating MALAT1. MeRIP-qPCR confirmed that RBM10 could inhibit the MALAT1 m6A methylation level by recruiting Methyltransferase Like 3 (METTL3). SIGNIFICANCE: The study suggests that RBM10, as an RNA-binding protein, may inhibit the m6A methylation of MALAT1 by recruiting METTL3 and affecting phosphorylation of the downstream PI3K/AKT/mTOR pathway by binding and regulating MALAT1, ultimately affecting the invasion and migration of NSCLC.


Subject(s)
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , RNA, Long Noncoding , Humans , Carcinoma, Non-Small-Cell Lung/pathology , Lung Neoplasms/pathology , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Cell Line, Tumor , TOR Serine-Threonine Kinases/metabolism , Methyltransferases/genetics , Methyltransferases/metabolism , RNA-Binding Proteins/genetics
20.
Nature ; 613(7943): 383-390, 2023 Jan.
Article in English | MEDLINE | ID: mdl-36599982

ABSTRACT

Specific, regulated modification of RNAs is important for proper gene expression1,2. tRNAs are rich with various chemical modifications that affect their stability and function3,4. 7-Methylguanosine (m7G) at tRNA position 46 is a conserved modification that modulates steady-state tRNA levels to affect cell growth5,6. The METTL1-WDR4 complex generates m7G46 in humans, and dysregulation of METTL1-WDR4 has been linked to brain malformation and multiple cancers7-22. Here we show how METTL1 and WDR4 cooperate to recognize RNA substrates and catalyse methylation. A crystal structure of METTL1-WDR4 and cryo-electron microscopy structures of METTL1-WDR4-tRNA show that the composite protein surface recognizes the tRNA elbow through shape complementarity. The cryo-electron microscopy structures of METTL1-WDR4-tRNA with S-adenosylmethionine or S-adenosylhomocysteine along with METTL1 crystal structures provide additional insights into the catalytic mechanism by revealing the active site in multiple states. The METTL1 N terminus couples cofactor binding with conformational changes in the tRNA, the catalytic loop and the WDR4 C terminus, acting as the switch to activate m7G methylation. Thus, our structural models explain how post-translational modifications of the METTL1 N terminus can regulate methylation. Together, our work elucidates the core and regulatory mechanisms underlying m7G modification by METTL1, providing the framework to understand its contribution to biology and disease.


Subject(s)
Cryoelectron Microscopy , GTP-Binding Proteins , Methylation , Methyltransferases , RNA Processing, Post-Transcriptional , RNA, Transfer , Humans , Catalytic Domain , Crystallography, X-Ray , GTP-Binding Proteins/chemistry , GTP-Binding Proteins/metabolism , GTP-Binding Proteins/ultrastructure , Methyltransferases/chemistry , Methyltransferases/metabolism , Methyltransferases/ultrastructure , RNA, Transfer/chemistry , RNA, Transfer/metabolism , RNA, Transfer/ultrastructure , S-Adenosylhomocysteine/metabolism , S-Adenosylmethionine/metabolism , Substrate Specificity , Biocatalysis
SELECTION OF CITATIONS
SEARCH DETAIL