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M5C methyltransferase NSUN3 regulates variant gene expression and development in blood-stage Plasmodium falciparum [RIP-seq]

ABSTRACT: m5C methyltransferase NSUN3 regulates variant gene expression and development in blood-stage Plasmodium falciparum [RIP-seq]

PROVIDER: PRJNA1139780 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR29967051_1.fastq.gz	Fastqsanger.gz
	SRR29967051_2.fastq.gz	Fastqsanger.gz

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m5C methyltransferase NSUN3 regulates variant gene expression and development in blood-stage Plasmodium falciparum [RNA-seq]

Project description:m5C methyltransferase NSUN3 regulates variant gene expression and development in blood-stage Plasmodium falciparum [RNA-seq]

| PRJNA1139782 | ENA

m5C methyltransferase NSUN3 regulates variant gene expression and development in blood-stage Plasmodium falciparum [RIP-seq]

Project description:C5 methylation of cytosine is a relatively abundant post-transcriptional modification in eukaryotes, and the NSUN family has been identified as an important executor of m5C modification. Currently, little is known about this family in Plasmodium spp. Therefore, we constructed a pfnsun3 gene knockdown strain and the knockdown efficiency was confirmed through growth curves and western blot experiments. The target genes of PfNSUN3 was obtained by RNA immunoprecipitation and high-throughput transcriptome sequencing experiments. Our data reveal that pfnsun3 is an indispensable post-transcriptional RNA modification in regulating variant gene expression in Plasmodium falciparum.

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Study on the Mechanism of m5C Methyltransferase NSUN3 Remodeling the Glioma Immune Microenvironment by Driving Tunneling Nanotube-Mediated Mitochondrial Transfer via tRNA Modification

Project description:Glioma is characterized by an immunosuppressive tumor microenvironment (TME) that promotes immune escape and therapeutic resistance. Mitochondrial transfer mediated by tunneling nanotubes (TNTs) is critical for tumor-stroma crosstalk, yet the underlying epigenetic regulatory mechanism remains unclear. Here, we report that the RNA m5C methyltransferase NSUN3 drives TNT formation and intercellular mitochondrial transfer in glioma by catalyzing m5C modification on tRNAs. Using tRNA-specific bisulfite sequencing (tRNA-BSseq) on human glioma cells, we systematically profiled tRNA m5C methylation landscapes in NSUN3-overexpressing and control groups. We identified NSUN3-dependent m5C sites on tRNAs that modulate translation efficiency of key factors involved in TNT assembly and mitochondrial transport. Functional assays demonstrated that NSUN3-mediated tRNA m5C modification enhances TNT formation, facilitates mitochondrial transfer from stromal cells to glioma cells, and remodels the immune microenvironment by suppressing anti-tumor immunity and promoting immunosuppressive cell infiltration. Our study reveals a novel epitranscriptomic mechanism linking tRNA m5C methylation to intercellular organelle transfer and immune regulation in glioma, providing potential targets for glioma immunotherapy.

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m5C methyltransferase NSUN3 regulates variant gene expression and development in blood-stage Plasmodium falciparum [RNA-seq]

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Study on the Mechanism of m5C Methyltransferase NSUN3 Remodeling the Glioma Immune Microenvironment by Driving Tunneling Nanotube-Mediated Mitochondrial Transfer via tRNA Modification [RNA-Seq]

Project description:Glioma is the most common and aggressive malignant tumor in the central nervous system, and its immunosuppressive microenvironment is closely associated with therapeutic resistance and poor prognosis. NSUN3, an m5C methyltransferase, catalyzes m5C modification on tRNA and regulates RNA translation and cellular metabolism. However, the role and transcriptomic mechanism of NSUN3 in remodeling the glioma immune microenvironment remain unclear. In this study, mRNA sequencing was performed on NSUN3-knockdown glioma cells and control cells to identify differentially expressed mRNAs and related signaling pathways. Total RNA was extracted, and mRNA was enriched for library construction and high-throughput sequencing. Bioinformatics analyses including quality control, read mapping, gene quantification, differential expression screening, and functional enrichment (GO, KEGG, GSEA) were conducted. We identified a series of differentially expressed mRNAs associated with tRNA modification, mitochondrial transfer, tunneling nanotube formation, and immune regulation. These mRNAs were significantly enriched in immune response, cellular metabolism, and tumor progression pathways. Our transcriptomic data provide a comprehensive resource for understanding the mechanism by which NSUN3 drives tunneling nanotube-mediated mitochondrial transfer and remodels the glioma immune microenvironment, and offer potential targets for glioma immunotherapy.

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Project description:Methylation of carbon 5 in cytosine (5-methylcytosine; m5C) is a well-characterized DNA modification, and is also predominantly reported in highly abundant noncoding RNAs, such as rRNA and tRNA, in both prokaryotes and eukaryotes. However, the distribution and biological functions of m5C in plant mRNAs remain largely unknown. Here we develop an m5C RNA immunoprecipitation followed by deep sequencing approach (m5C-RIP-seq) to achieve transcriptome-wide profiling of RNA m5C in Arabidopsis thaliana. Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and dot blot analyses reveal a dynamic pattern of m5C mRNA modification in various tissues and at different developmental stages. m5C-RIP-seq analysis identifies 6,045 putative m5C peaks in 4,465 expressed genes in young seedlings. m5C is enriched in coding sequences with two peaks located immediately after start codons and before stop codons, and is associated with mRNAs with low translation activity. We further show that a RNA (cytosine-5)-methyltransferase, tRNA specific methyltransferase 4B (TRM4B), exhibits the m5C mRNA methyltransferase activity. Mutations in TRM4B display defects in root development and decreased m5C levels in root mRNA. Furthermore, TRM4B affects transcript levels of the genes involved in root development, which is positively correlated with their mRNA stability and m5C levels. Our results suggest that m5C in mRNA is a new epitranscriptome marker widely distributed in plant genes, and that regulation of this modification is an integral part of gene regulatory networks underlying plant development.

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The RNA M5C Methyltransferase NSUN2 Promotes Progression of Hepatocellular Carcinoma by Enhancing PKM2-Mediated Glycolysis

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Project description:Aberrant post-transcriptional methylation is implicated in a wide range of human diseases, yet the specific enzymes responsible for site- and substrate-specific methylation remain largely unknown. Here, we use our recently developed catalysis-dependent RIP sequencing approach (miCLIP) and RNA bisulfite sequencing to map C5-methylcytosine (m5C) in the human transcriptome. We identified two novel m5C methylases NSun3 and NSun6, both of which have strong substrate specificity. In contrast to the previously characterized NSun2, which displays some preference to methylating transfer RNAs (tRNA), NSun6 predominantly targeted 3’ UTRs of messenger RNAs (mRNA), and NSun3 mainly methylated mitochondrial RNAs (mtRNA). Consistent with the miCLIP-predicted RNA target specificity, whole exome sequencing identified NSUN3 loss-of-function mutations in a patient presenting with combined respiratory chain complex deficiency. Functional studies of the patient fibroblast cell line revealed that loss of the NSun3 protein resulted in severe mitochondrial translation defects, which were rescued by expression of wild-type NSun3. In summary, our results reveal how highly conserved NSun m5C methylases partition their substrate specificities to remodel the sequences of particular ribonucleotide classes.

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