TGF-β-SMAD signaling pathway plays an important role in the progression of various cancers. However, posttranscriptional regulation such as N6-methyladenosine (m6A) of TGF-β-SMAD signaling axis remains incompletely understood. Here, we reveal that insulin like growth factor 2 mRNA binding protein 2 (IGF2BP2) is low expression as well as associated with poor prognosis in clear cell renal cell carcinoma (ccRCC) patients and inhibits proliferation as well as promotes metastasis of ccRCC cells. Mechanistically, IGF2BP2 systematically regulates TGF-β-SMAD signaling family, including TGF-β1/2, TGF-βR1/2 and SMAD2/3/4, through mediating their mRNA stability in an m6A-dependent manner. Furthermore, the functional effects of IGF2BP2 on ccRCC cells is mediated by TGF-β-SMAD signaling downstream effector SMAD4, which is identified three m6A sites in 5\'UTR and CDS. Our study establishes IGF2BP2-TGF-β-SMAD axis as a new regulatory effector in ccRCC, providing new insights for developing novel therapeutic strategies.

Targeting RNA m6A marks in apoptosis-related transcripts holds promise for RNA therapeutics. However, pathway-specific RNA m6A sites on pro- or antiapoptotic transcripts have not been fully unveiled, let alone characterized. This article summarizes the current knowledge and gaps in the cellular response modulated by apoptotic stimulus-specific RNA m6A marks.

BACKGROUND: The regulatory mechanisms of RNA methylation during the processes of osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) have yet to be fully understood. The objective of our study was to analyze and validate the contribution of RNA methylation regulators to the mechanisms underlying the osteogenic and adipogenic differentiation of rat BMSCs.
METHODS: We downloaded the GSE186026 from the Gene Expression Omnibus (GEO). Differentially expressed genes (DEGs) were screened using the DESeq2 package in R software (version 3.6.3). A total of 50 RNA methylation genes obtained from literature review and summary were intersected with the previous DEGs to obtain RNA methylation genes, which have different expressions (RM-DEGs). Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were utilized to reveal the functional enrichment. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to validate RM-DEGs. Protein-protein interaction network (PPI) analysis and visual analysis were performed using STRING and Cytoscape. RM-DEGs regulatory network was constructed to analyze the top 10 hub genes. The relationship between RM-DEGs, some enriched GO and pathways was also been analyzed. The miRNAs and RM-DEGs regulatory networks were established by using miRWalk and TargetScan.
RESULTS: As part of our research, we detected varying levels of expression for m6A regulators Mettl3 and Rbm15, as well as m7G regulators Mettl1 and Wdr4, in relation to osteogenic differentiation, along with m6A regulator Fmr1 in adipogenic differentiation. The protein-protein interaction (PPI) networks were constructed for 49 differentially expressed genes (DEGs) related to RNA methylation during the process of osteogenic differentiation, and 13 DEGs for adipogenic differentiation. Moreover, top10 hub genes were calculated. In osteogenic differentiation, Mettl3 regulated the Wnt pathway and Hippo pathway by regulating Smad3, Rbm15 regulated the Notch pathway by Notch1, Mettl1 regulated the PI3K-Akt pathway by Gnb4. In adipogenic differentiation, Fmr1 regulated the PI3K-Akt pathway by Egfr. M6A methylation sites of Smad3, Notch1 and Gnb4 were predicted, and the results showed that all three genes were possibly methylated by m6A, and more than 9 sites per gene were possibly methylated. Finally, we constructed the regulatory networks of Mettl3, Rbm15, Mettl1, and Wdr4 and 109 miRNAs in osteogenic differentiation, Fmr1 and 118 miRNAs in adipogenic differentiation.
CONCLUSIONS: Mettl3(m6A), Rbm15(m6A), Wdr4 and Mettl1(m7G) were differentially expressed in osteogenic differentiation, while Fmr1(m6A) was differentially expressed in adipogenic differentiation. These findings offered potential candidates for further research on the involvement of RNA methylation in the osteogenic and adipogenic differentiation of BMSCs.

OBJECTIVE: This study will explore the function of WTAP, the critical segment of m6A methyltransferase complex, in UC and its regulation on immune response.
METHODS: The expression levels of key proteins were detected in colon tissues which were derived from UC patients and mice. Macrophage polarization and CD4+ T cell infiltration were detected by flow cytometry and IF staining. ELISA assay was utilized to analyze the level of the inflammatory cytokines. m6A-RIP-PCR, actinomycin D test, and RIP assays were utilized to detect the m6A level, stability, and bound proteins of CES2 mRNA. A dual luciferase reporter assay was conducted to confirm the transcriptional interactions between genes. A co-culture system of intestinal epithelium-like organs was constructed to detect the primary mouse intestinal epithelial cells (PMIEC) differentiation. The interaction between proteins was detected via Co-IP assay.
RESULTS: The expression of WTAP and CES2 in UC tissues was increased and decreased, respectively. Knockdown of WTAP inhibited the progression of UC in mice by inhibiting M1 macrophage polarization and CD4+ T cell infiltration. WTAP combined YTHDF2 to promote the m6A modification of CES2 mRNA and inhibited its expression. CES2 co-expressed with EPHX2 and overexpression of CES2 promoted the differentiation of PMIEC. The inhibitory effect of WTAP knockdown on the progress of UC was partially abrogated by CES2 knockdown.
CONCLUSIONS: WTAP/YTHDF2 silences CES2 by promoting its m6A modification and then promotes the progression of UC. WTAP could be a promoting therapy target of UC.

N6-methyladenosine (m6A) methylation is the most prevalent post-transcriptional RNA modification in eukaryotic organisms, but its roles in the regulation of physiological resistance of marine crustaceans to heavy metal pollutants are poorly understood. In this study, the transcriptome-wide m6A RNA methylation profiles and dynamic m6A changes induced by acute Cd2+ exposure in the the pacific whiteleg shrimp Litopenaeus vannamei were comprehensively analyzed. Cd2+ toxicity caused a significant reduction in global RNA m6A methylation level, with major m6A regulators including the m6A methyltransferase METTL3 and the m6A binding protein YTHDF2 showing declined expression. Totally, 11,467 m6A methylation peaks from 6415 genes and 17,291 peaks within 7855 genes were identified from the Cd2+ exposure group and the control group, respectively. These m6A peaks were predominantly enriched in the 3\' untranslated region (UTR) and around the start codon region of the transcripts. 7132 differentially expressed genes (DEGs) and 7382 differentially m6A-methylated genes (DMGs) were identified. 3186 genes showed significant changes in both gene expression and m6A methylation levels upon cadmium exposure, and they were related to a variety of biological processes and gene pathways. Notably, an array of genes associated with antioxidation homeostasis, transmembrane transporter activity and intracellular detoxification processes were significantly enriched, demonstrating that m6A modification may mediate the physiological responses of shrimp to cadmium toxicity via regulating ROS balance, Cd2+ transport and toxicity mitigation. The study would contribute to a deeper understanding of the evolutionary and functional significance of m6A methylation to the physiological resilience of decapod crustaceans to heavy metal toxicants.

PM2.5 pollution has been associated with the incidence of lung cancer, but the underlying mechanism is still unclear. PIWI-interacting RNAs (piRNAs), initially identified in germline cells, have emerged as a novel class of small non-coding RNAs (26 - 32 nucleotides) with diverse functions in various diseases, including cancer. However, the role and mechanism of piRNAs in the development of PM2.5-induced lung cancer remain to be clarified. In the presented study, we used a PM2.5-induced malignant transformation cell model to analyze the change of piRNA profiles. Among the disturbed piRNAs, piR-27222 was identified as an oncogene that inhibited cell death in a m6A-dependent manner. Mechanistically, we found that piR-27222 could deubiquitinate and stabilize eIF4B by directly binding to eIF4B and reducing its interaction with PARK2. The enhanced expression of eIF4B, in turn, promoted the expression of WTAP, leading to increased m6A modification in the Casp8 transcript. Consequently, the stability of Casp8 transcripts was reduced, rendering lung cancer cells resistant to PANoptosis. Collectively, our findings reveal that PM2.5 exposure up-regulated piR-27222 expression, which could affect EIF4B/WTAP/m6A axis, thereby inhibiting PANoptosis of cells and promoting lung cancer. Our study provides new insights into understanding the epigenetic mechanisms underlining PM2.5-induced lung cancer.

N6-methyladenosine (m6A), the most abundant modification in mRNAs, affects the fate of the modified RNAs at the post-transcriptional level and participants in various biological and pathological processes. Increasing evidence shows that m6A modification plays a role in the progression of many malignancies, including colorectal cancer (CRC). As the only catalytic subunit in methyltransferase complex, methyltransferase-like 3 (METTL3) is essential to the performance of m6A modification. It has been found that METTL3 is associated with the prognosis of CRC and significantly influences various aspects of CRC, such as cell proliferation, invasion, migration, metastasis, metabolism, tumor microcirculation, tumor microenvironment, and drug resistance. The relationship between METTL3 and gut-microbiota is also involved into the progression of CRC. Furthermore, METTL3 might be a viable target for CRC treatment to prolong survival. In this review, we comprehensively summarize the function of METTL3 in CRC and the underlying molecular mechanisms. We aim to deepen understanding and offer new ideas for diagnostic biomarkers and therapeutic targets for colorectal cancer.

Genomic instability poses a formidable threat to cellular vitality and wellbeing, prompting cells to deploy an intricate DNA damage response (DDR) mechanism. Recent evidence has suggested that RNA is intricately linked to the DDR by serving as template, scaffold, or regulator during the repair of DNA damage. Additionally, RNA molecules undergo modifications, contributing to the epitranscriptome, a dynamic regulatory layer influencing cellular responses to genotoxic stress. The intricate interplay between RNA and the DDR sheds new light on how the RNA epigenome contributes to the maintenance of genomic integrity and ultimately shapes the fate of damaged cells.

The imbalance of vascular endothelial cell homeostasis is the key mechanism for the progression of many vascular diseases. RNA modification, particularly N6-Methyladenosine (m6A), plays important function in numerous biological processes. Nevertheless, the regulatory function of m6A RNA methylation in endothelial dysfunction remains insufficiently characterized. In this study, we established that the m6A methyltransferase METTL3 is critical for regulating endothelial function. Functionally, depletion of METTL3 results in decreased endothelial cells proliferation, survival and inflammatory response. Conversely, overexpression of METTL3 elicited the opposite effects. Mechanistically, MeRIP-seq identified that METTL3 catalyzed m6A modification of TRAF1 mRNA and enhanced TRAF1 translation, thereby up-regulation of TRAF1 protein. Over-expression of TRAF1 successfully rescued the inhibition of proliferation and adhesion of endothelial cells due to METTL3 knockdown. Additionally, m6A methylation-mediated TRAF1 expression can be reversed by the demethylase ALKBH5. Knockdown of ALKBH5 upregulated the level of m6A and protein level of TRAF1, and also increased endothelial cells adhesion and inflammatory response. Collectively, our findings suggest that METTL3 regulates vascular endothelium homeostasis through TRAF1 m6A modification, suggesting that targeting the METTL3-m6A-TRAF1 axis may hold therapeutic potential for patients with vascular diseases.

Blood development and regeneration require rapid turnover of cells, and ribonucleic acid (RNA) modifications play a key role in it via regulating stemness and cell fate regulation. RNA modifications affect gene activity via posttranscriptional and translation-mediated mechanisms. Diverse molecular players involved in RNA-modification processes are abundantly expressed by hematopoietic stem cells and lineages. Close to 150 RNA chemical modifications have been reported, but only N6-methyl adenosine (m6A), inosine (I), pseudouridine (Ψ), and m1A-a handful-have been studied in-cell fate regulation. The role of RNA modification in blood diseases and disorders is an emerging field and offers potential for therapeutic interventions. Knowledge of RNA-modification and enzymatic activities could be used to design therapies in the future. Here, we summarized the recent advances in RNA modification and the epitranscriptome field and discussed their regulation of blood development and regeneration.