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Virulence Dec 2022infects various crops with aflatoxins, and leads to aspergillosis opportunistically. Though H3K36 methylation plays an important role in fungal toxin metabolism and...
infects various crops with aflatoxins, and leads to aspergillosis opportunistically. Though H3K36 methylation plays an important role in fungal toxin metabolism and virulence, no data about the biological function of H3K36 methylation in virulence has been reported. Our study showed that the Set2 histone methyltransferase family, AshA and SetB, involves in morphogenesis and mycotoxin anabolism by regulating related transcriptional factors, and they are important for fungal virulence to crops and animals. Western-blotting and double deletion analysis revealed that AshA mainly regulates H3K36me2, whereas SetB is mainly responsible for H3K36me3 in the nucleus. By construction of domain deletion strain and point mutation strains by homologous recombination, the study revealed that SET domain is indispensable in mycotoxin anabolism and virulence of , and N455 and V457 in it are the key amino acid residues. ChIP analysis inferred that the methyltransferase family controls fungal reproduction and regulates the production of AFB1 by directly regulating the production of the transcriptional factor genes, including , , and amylase, through H3K36 trimethylation in their chromatin fragments, based on which this study proposed that, by H3K36 trimethylation, this methyltransferase family controls AFB1 anabolism through transcriptional level and substrate utilization level. This study illuminates the epigenetic mechanism of the Set2 family in regulating fungal virulence and mycotoxin production, and provides new targets for controlling the virulence of the fungus .The methylation of H3K36 plays an important role in the fungal secondary metabolism and virulence, but no data about the regulatory mechanism of H3K36 methylation in the virulence of have been reported. Our study revealed that, in the histone methyltransferase Set2 family, AshA mainly catalyzes H3K36me2, and involves in the methylation of H3K36me1, and SetB mainly catalyzes H3K36me3 and H3K36me1. Through domain deletion and point mutation analysis, this study also revealed that the SET domain was critical for the normal biological function of the Set2 family and that N455 and V457 in the domain were critical for AshA. By ChIP-seq and ChIP-qPCR analysis, H3K36 was found to be trimethylation modified in the promotors and ORF positions of , and the amylase gene (AFLA_084340), and further qRT-PCR results showed that these methylation modifications regulate the expression levels of these genes. According to the results of ChIP-seq analysis, we proposed that, by H3K36 trimethylation, this methyltransferase family controls the metabolism of mycotoxin through transcriptional level and substrate utilization level. All the results from this study showed that Set2 family is essential for fungal secondary metabolism and virulence, which lays a theoretical groundwork in the early prevention and treatment of pollution, and also provides an effective strategy to fight against other pathogenic fungi.
Topics: Amylases; Animals; Aspergillus flavus; Histone Methyltransferases; Histones; Methylation; Methyltransferases; Mycotoxins; Secondary Metabolism; Virulence
PubMed: 35943142
DOI: 10.1080/21505594.2022.2101218 -
Molecular Plant Jul 2022Maintaining genomic integrity and stability is particularly important for stem cells, which are at the top of the cell lineage origin. Here, we discovered that the...
Maintaining genomic integrity and stability is particularly important for stem cells, which are at the top of the cell lineage origin. Here, we discovered that the plant-specific histone methyltransferase SUVR2 maintains the genome integrity of the root tip stem cells through chromatin remodeling and liquid-liquid phase separation (LLPS) when facing DNA double-strand breaks (DSBs). The histone methyltransferase SUVR2 (MtSUVR2) has histone methyltransferase activity and catalyzes the conversion of histone H3 lysine 9 monomethylation (H3K9me1) to H3K9me2/3 in vitro and in Medicago truncatula. Under DNA damage, the proportion of heterochromatin decreased and the level of DSB damage marker γ-H2AX increased in suvr2 mutants, indicating that MtSUVR2 promotes the compaction of the chromatin structure through H3K9 methylation modification to protect DNA from damage. Interestingly, MtSUVR2 was induced by DSBs to phase separate and form droplets to localize at the damage sites, and this was confirmed by immunofluorescence and fluorescence recovery after photobleaching experiments. The IDR1 and low-complexity domain regions of MtSUVR2 determined its phase separation in the nucleus, whereas the IDR2 region determined the interaction with the homologous recombinase MtRAD51. Furthermore, we found that MtSUVR2 drove the phase separation of MtRAD51 to form "DNA repair bodies," which could enhance the stability of MtRAD51 proteins to facilitate error-free homologous recombination repair of stem cells. Taken together, our study reveals that chromatin remodeling-associated proteins participate in DNA repair through LLPS.
Topics: Chromatin; Chromatin Assembly and Disassembly; DNA Breaks, Double-Stranded; DNA Repair; Histone Methyltransferases; Histones; Methyltransferases
PubMed: 35610973
DOI: 10.1016/j.molp.2022.05.007 -
Antiviral Research Oct 2008Many flaviviruses are significant human pathogens. No effective antiviral therapy is currently available for treatment of flavivirus infections. Development of antiviral... (Review)
Review
Many flaviviruses are significant human pathogens. No effective antiviral therapy is currently available for treatment of flavivirus infections. Development of antiviral treatment against these viruses is urgently needed. The flavivirus methyltransferase (MTase) responsible for N-7 and 2'-O methylation of the viral RNA cap has recently been mapped to the N-terminal region of nonstructural protein 5. Structural and functional studies suggest that the MTase represents a novel antiviral target. Here we review current understanding of flavivirus RNA cap methylation and its implications for development of antivirals. The 5' end of the flavivirus plus-strand RNA genome contains a type 1 cap structure (m(7)GpppAmG). Flaviviruses encode a single MTase domain that catalyzes two sequential methylations of the viral RNA cap, GpppA-RNA-->m(7)GpppA-RNA-->m(7)GpppAm-RNA, using S-adenosyl-L-methionine (SAM) as the methyl donor. The two reactions require different viral RNA elements and distinct biochemical assay conditions. Despite exhibiting two distinct methylation activities, flavivirus MTase contains a single binding site for SAM in its crystal structure. Therefore, substrate GpppA-RNA must be re-positioned to accept the N-7 and 2'-O methyl groups from SAM during the two methylation reactions. Structure-guided mutagenesis studies indeed revealed two distinct sets of amino acids on the enzyme surface that are specifically required for N-7 and 2'-O methylation. In the context of virus, West Nile viruses (WNVs) defective in N-7 methylation are non-replicative; however, WNVs defective in 2'-O methylation are attenuated and can protect mice from subsequent wild-type WNV challenge. Collectively, the results demonstrate that the N-7 MTase represents a novel target for flavivirus therapy.
Topics: Amino Acid Sequence; Antiviral Agents; Flavivirus; Humans; Methylation; Methyltransferases; Models, Molecular; Molecular Sequence Data; RNA Caps; RNA, Viral; Virus Replication
PubMed: 18571739
DOI: 10.1016/j.antiviral.2008.05.003 -
Biomolecules Jan 2017Post-transcriptional and post-translational modifications are very important for the control and optimal efficiency of messenger RNA (mRNA) translation. Among these,... (Review)
Review
Post-transcriptional and post-translational modifications are very important for the control and optimal efficiency of messenger RNA (mRNA) translation. Among these, methylation is the most widespread modification, as it is found in all domains of life. These methyl groups can be grafted either on nucleic acids (transfer RNA (tRNA), ribosomal RNA (rRNA), mRNA, etc.) or on protein translation factors. This review focuses on Trm112, a small protein interacting with and activating at least four different eukaryotic methyltransferase (MTase) enzymes modifying factors involved in translation. The Trm112-Trm9 and Trm112-Trm11 complexes modify tRNAs, while the Trm112-Mtq2 complex targets translation termination factor eRF1, which is a tRNA mimic. The last complex formed between Trm112 and Bud23 proteins modifies 18S rRNA and participates in the 40S biogenesis pathway. In this review, we present the functions of these eukaryotic Trm112-MTase complexes, the molecular bases responsible for complex formation and substrate recognition, as well as their implications in human diseases. Moreover, as Trm112 orthologs are found in bacterial and archaeal genomes, the conservation of this Trm112 network beyond eukaryotic organisms is also discussed.
Topics: Amino Acid Sequence; Eukaryotic Cells; Methyltransferases; Models, Molecular; Prokaryotic Cells; Protein Biosynthesis
PubMed: 28134793
DOI: 10.3390/biom7010007 -
Current Medicinal Chemistry 2010DNA methylation is an epigenetic event involved in a variety array of processes that may be the foundation of genetic phenomena and diseases. DNA methyltransferase is a... (Review)
Review
DNA methylation is an epigenetic event involved in a variety array of processes that may be the foundation of genetic phenomena and diseases. DNA methyltransferase is a key enzyme for cytosine methylation in DNA, and can be divided into two functional families (Dnmt1 and Dnmt3) in mammals. All mammalian DNA methyltransferases are encoded by their own single gene, and consisted of catalytic and regulatory regions (except Dnmt2). Via interactions between functional domains in the regulatory or catalytic regions and other adaptors or cofactors, DNA methyltransferases can be localized at selective areas (specific DNA/nucleotide sequence) and linked to specific chromosome status (euchromatin/heterochromatin, various histone modification status). With assistance from UHRF1 and Dnmt3L or other factors in Dnmt1 and Dnmt3a/Dnmt3b, mammalian DNA methyltransferases can be recruited, and then specifically bind to hemimethylated and unmethylated double-stranded DNA sequence to maintain and de novo setup patterns for DNA methylation. Complicated enzymatic steps catalyzed by DNA methyltransferases include methyl group transferred from cofactor Ado-Met to C5 position of the flipped-out cytosine in targeted DNA duplex. In the light of the fact that different DNA methyltransferases are divergent in both structures and functions, and use unique reprogrammed or distorted routines in development of diseases, design of new drugs targeting specific mammalian DNA methyltransferases or their adaptors in the control of key steps in either maintenance or de novo DNA methylation processes will contribute to individually treating diseases related to DNA methyltransferases.
Topics: Animals; DNA; DNA (Cytosine-5-)-Methyltransferase 1; DNA (Cytosine-5-)-Methyltransferases; DNA Methylation; Epigenesis, Genetic; Humans; Mammals; Molecular Targeted Therapy; Regulatory Sequences, Nucleic Acid; DNA Methyltransferase 3B
PubMed: 20939822
DOI: 10.2174/092986710793205372 -
Protein Science : a Publication of the... Sep 2018Protein, DNA, and RNA methyltransferases have an ever-expanding list of novel substrates and catalytic activities. Even within families and between homologs, it is...
Protein, DNA, and RNA methyltransferases have an ever-expanding list of novel substrates and catalytic activities. Even within families and between homologs, it is becoming clear the intricacies of methyltransferase specificity and regulation are far more diverse than originally thought. In addition to specific substrates and distinct methylation levels, methyltransferase activity can be altered by complex formation with close homologs. We work with the N-terminal methyltransferase homologs NRMT1 and NRMT2. NRMT1 is a ubiquitously expressed distributive trimethylase. NRMT2 is a monomethylase expressed at low levels in a tissue-specific manner. They are both nuclear methyltransferases with overlapping consensus sequences but have distinct enzymatic activities and tissue expression patterns. Co-expression with NRMT2 increases the trimethylation rate of NRMT1, and here we aim to understand how this occurs. We use analytical ultracentrifugation to show that while NRMT1 primarily exists as a dimer and NRMT2 as a monomer, when co-expressed they form a heterotrimer. We use co-immunoprecipitation and molecular modeling to demonstrate in vivo binding and map areas of interaction. While overexpression of NRMT2 increases the half-life of NRMT1, the converse is not true, indicating that NRMT2 may be increasing NRMT1 activity by stabilizing the enzyme. Accordingly, the catalytic activity of NRMT2 is not needed to increase NRMT1 activity or increase its affinity for less preferred substrates. Monomethylation can also not rescue phenotypes seen with loss of trimethylation. Taken together, these data support a model where NRMT2 expression activates NRMT1 activity, not through priming, but by increasing its stability and substrate affinity.
Topics: Biocatalysis; Humans; Isoenzymes; Methyltransferases; Models, Molecular
PubMed: 30151928
DOI: 10.1002/pro.3456 -
The Plant Cell May 2023Addition of N6-methyladenosine (m6A), the most prevalent internal mRNA modification in eukaryotes, is catalyzed by an evolutionarily conserved m6A methyltransferase...
Addition of N6-methyladenosine (m6A), the most prevalent internal mRNA modification in eukaryotes, is catalyzed by an evolutionarily conserved m6A methyltransferase complex. In the model plant Arabidopsis thaliana, the m6A methyltransferase complex is composed of 2 core methyltransferases, mRNA adenosine methylase (MTA) and MTB, and several accessory subunits such as FK506-BINDING PROTEIN 12 KD INTERACTING PROTEIN 37KD (FIP37), VIRILIZER (VIR), and HAKAI. It is yet largely unknown whether these accessory subunits influence the functions of MTA and MTB. Herein, I reveal that FIP37 and VIR are indispensable for stabilizing the methyltransferases MTA and MTB, thus functioning as key subunits to maintain the functionality of the m6A methyltransferase complex. Furthermore, VIR affects FIP37 and HAKAI protein accumulation, while MTA and MTB mutually influence each other. In contrast, HAKAI has little effect on protein abundance or localization of MTA, MTB, and FIP37. These findings uncover unique functional interdependence at the post-translational level among individual components in the Arabidopsis m6A methyltransferase complex, suggesting that maintenance of protein homeostasis among various subunits of the m6A methyltransferase complex is essential for maintaining the protein stoichiometry required for the proper function of the m6A methyltransferase complex in m6A deposition in plants.
Topics: Arabidopsis; Methyltransferases; RNA, Messenger; Adenosine
PubMed: 36890720
DOI: 10.1093/plcell/koad070 -
Frontiers in Immunology 2022The aim of the current study was to investigate the contributing role of gene variation and transcription levels among the m6A methyltransferases METTL3, METTL14, and...
OBJECTIVE
The aim of the current study was to investigate the contributing role of gene variation and transcription levels among the m6A methyltransferases METTL3, METTL14, and WTAP in pulmonary tuberculosis (PTB).
METHODS
A case-control study including 461 PTB patients and 467 normal controls was designed for genotyping. Three SNPs in (rs1061027, rs1139130, rs1061026), three SNPs in (rs62328061, rs4834698, rs1064034), and two SNPs in (rs1853259, rs11752345) were genotyped the SNPscan™ technique. , , and transcription levels were determined in 78 PTB patients and 86 controls quantitative real-time reverse-transcription PCR.
RESULTS
Frequencies of the rs62328061 GG genotype, rs11752345 CT genotype, and T allele were significantly increased in PTB patients compared to controls. An increased risk of rs62328061 was detected in a recessive model, and a decreased risk of rs11752345 was detected in a dominant model in the PTB group. gene variation was not associated with PTB risk. The rs1139130 GG genotype was significantly increased with drug resistance, and the G allele was significantly decreased with drug-induced liver injury in PTB patients. A reduced frequency of the rs62328061 G allele was associated with leukopenia, a reduced frequency of the rs11752345 T allele was associated with sputum smear positivity, and a higher frequency of the rs4834698 TC genotype was evident in PTB patients with hypoproteinemia. Compared to controls, , , and transcription levels in PTB patients were significantly decreased, and the level of WTAP was increased in PTB patients with drug resistance. METTL3 level was negatively associated with erythrocyte sedimentation rate and aspartate aminotransferase, and METTL14 level was negatively correlated with alanine aminotransferase and aspartate aminotransferase.
CONCLUSION
rs62328061 and rs11752345 variants were associated with the genetic background of PTB, and METTL3, METTL14, and WTAP levels were abnormally decreased, suggesting that these m6A methyltransferases may play important roles in PTB.
Topics: Humans; Case-Control Studies; RNA, Messenger; Methyltransferases; RNA Splicing Factors; Cell Cycle Proteins
PubMed: 36569923
DOI: 10.3389/fimmu.2022.992628 -
Journal of Virology Mar 2017The Zika virus (ZIKV) is the causal agent of neurological disorders like microcephaly in newborns or Guillain-Barre syndrome. Its NS5 protein embeds a methyltransferase...
The Zika virus (ZIKV) is the causal agent of neurological disorders like microcephaly in newborns or Guillain-Barre syndrome. Its NS5 protein embeds a methyltransferase (MTase) domain involved in the formation of the viral mRNA cap. We investigated the structural and functional properties of the ZIKV MTase. We show that the ZIKV MTase can methylate RNA cap structures at the N-7 position of the cap, and at the 2'-O position on the ribose of the first nucleotide, yielding a cap-1 structure. In addition, the ZIKV MTase methylates the ribose 2'-O position of internal adenosines of RNA substrates. The crystal structure of the ZIKV MTase determined at a 2.01-Å resolution reveals a crystallographic homodimer. One chain is bound to the methyl donor (-adenosyl-l-methionine [SAM]) and shows a high structural similarity to the dengue virus (DENV) MTase. The second chain lacks SAM and displays conformational changes in the αX α-helix contributing to the SAM and RNA binding. These conformational modifications reveal a possible molecular mechanism of the enzymatic turnover involving a conserved Ser/Arg motif. In the second chain, the SAM binding site accommodates a sulfate close to a glycerol that could serve as a basis for structure-based drug design. In addition, compounds known to inhibit the DENV MTase show similar inhibition potency on the ZIKV MTase. Altogether these results contribute to a better understanding of the ZIKV MTase, a central player in viral replication and host innate immune response, and lay the basis for the development of potential antiviral drugs. The Zika virus (ZIKV) is associated with microcephaly in newborns, and other neurological disorders such as Guillain-Barre syndrome. It is urgent to develop antiviral strategies inhibiting the viral replication. The ZIKV NS5 embeds a methyltransferase involved in the viral mRNA capping process, which is essential for viral replication and control of virus detection by innate immune mechanisms. We demonstrate that the ZIKV methyltransferase methylates the mRNA cap and adenosines located in RNA sequences. The structure of ZIKV methyltransferase shows high structural similarities to the dengue virus methyltransferase, but conformational specificities highlight the role of a conserved Ser/Arg motif, which participates in RNA and SAM recognition during the reaction turnover. In addition, the SAM binding site accommodates a sulfate and a glycerol, offering structural information to initiate structure-based drug design. Altogether, these results contribute to a better understanding of the methyltransferases, which are central players in the virus replication.
Topics: Allosteric Site; Antiviral Agents; Catalytic Domain; Crystallography, X-Ray; Drug Design; Escherichia coli; Hydrogen Bonding; Methyltransferases; Models, Molecular; Protein Binding; Viral Nonstructural Proteins; Zika Virus
PubMed: 28031359
DOI: 10.1128/JVI.02202-16 -
Nature Chemistry Nov 2022Microviridins and other ω-ester-linked peptides, collectively known as graspetides, are characterized by side-chain-side-chain linkages installed by ATP-grasp enzymes....
Microviridins and other ω-ester-linked peptides, collectively known as graspetides, are characterized by side-chain-side-chain linkages installed by ATP-grasp enzymes. Here we report the discovery of a family of graspetides, the gene clusters of which also encode an O-methyltransferase with homology to the protein repair catalyst protein L-isoaspartyl methyltransferase. Using heterologous expression, we produced fuscimiditide, a ribosomally synthesized and post-translationally modified peptide (RiPP). NMR analysis of fuscimiditide revealed that the peptide contains two ester cross-links forming a stem-loop macrocycle. Furthermore, an unusually stable aspartimide moiety is found within the loop macrocycle. We fully reconstituted fuscimiditide biosynthesis in vitro including formation of the ester and aspartimide moieties. The aspartimide moiety embedded in fuscimiditide hydrolyses regioselectively to isoaspartate. Surprisingly, this isoaspartate-containing peptide is also a substrate for the L-isoaspartyl methyltransferase homologue, thus driving any hydrolysis products back to the aspartimide form. Whereas an aspartimide is often considered a nuisance product in protein formulations, our data suggest that some RiPPs have aspartimide residues intentionally installed via enzymatic activity.
Topics: Isoaspartic Acid; Amino Acid Sequence; Protein D-Aspartate-L-Isoaspartate Methyltransferase; Peptides; Methyltransferases; Esters
PubMed: 35982233
DOI: 10.1038/s41557-022-01022-y