Crosstalk between N6-methyladenosine (m6A) modification and noncoding RNA in tumor microenvironment

<p>RNA N6-methyladenosine (m<sup>6</sup>A) modification in tumorigenesis and progression has been highlighted and discovered in recent years. However, the molecular and clinical implications of m<sup>6</sup>A modification in melanoma tumor microenvironment (TME) and immune infiltration remain largely unknown. Here, we utilized consensus molecular clustering with nonnegative matrix factorization based on the melanoma transcriptomic profiles of 23 m<sup>6</sup>A regulators to determine the m<sup>6</sup>A modification clusters and m<sup>6</sup>A-related gene signature. Three distinct m<sup>6</sup>A modification patterns (m<sup>6</sup>A-C1, C2, and C3), which are characterized by specific m<sup>6</sup>A regulator expression, survival outcomes, and biological pathways, were identified in more than 1,000 melanoma samples. The immune profile analyses showed that these three m<sup>6</sup>A modification subtypes were highly consistent with the three known immune phenotypes: immune-desert (C1), immune-excluded (C2), and immune-inflamed (C3). Tumor digital cytometry (CIBERSORT, ssGSEA) algorithm revealed an upregulated infiltration of CD8<sup>+</sup> T cell and NK cell in m<sup>6</sup>A-C3 subtype. An m<sup>6</sup>A scoring scheme calculated by principal component of m<sup>6</sup>A signatures stratified melanoma patients into high- and low-m<sup>6</sup>sig score subgroups; a high score was significantly associated with prolonged survival and enhanced immune infiltration. Furthermore, fewer somatic copy number alternations (SCNA) and PD-L1 expression were found in patients with high m<sup>6</sup>Sig score. In addition, patients with high m<sup>6</sup>Sig score demonstrated marked immune responses and durable clinical benefits in two independent immunotherapy cohorts. Overall, this study indicated that m<sup>6</sup>A modification is involved in melanoma tumor microenvironment immune regulation and contributes to formation of tumor immunogenicity. Comprehensive evaluation of the m<sup>6</sup>A modification pattern of individual tumors will provide more insights into molecular mechanisms of TME characterization and promote more effective personalized biotherapy strategies.</p>

BackgroundThe association involving N6-methyladenosine (m6A) modification, molecular subtype and specific immune cell group in tumor microenvironment has been the focus of recent studies. The underlying function of m6A modification in thyroid cancer (TC) remains elusive.MethodsThe m6A modification regulations, molecular character and tumor immune profile of 461 TC patients were explored and then the correlation between them were comprehensively evaluated. The m6Ascore was established using principal component analysis (PCA) to quantify the m6A pattern of individual TC patients. The prognostic significance of the m6Ascore was evaluated by multivariate Cox regression analysis.ResultsFour m6Aclusters (mc1, 2, 3, 4)—characterized by differences in extent of aneuploidy, expression of immunomodulatory genes, mRNA or lncRNA expression pattern and prognosis were identified. T Preliminary validation of m6Ascore was a potential independent prognostic factor of TC involving in mc3. Finally, the prognostic value of the m6Ascore and its association with copy number variation (CNV) and tumor immune microenvironment (TIME) of TC in mc3 were verified.ConclusionsThe correlation between m6A modification, the copy number burden and tumor immune landscape in TC was demonstrated. A m6Acluster-mc3 with low m6Ascore and high CNV molecular subtype was identified with poor clinical prognosis, low infiltrating immunocyte and weak effector T cell. A three-gene clinical prognosis model for TC based on 4 m6a cluster expression was established. Understanding of TIME is enhanced by comprehensive assessment of m6A patterns in individual TC patients and gives a new insight toward improved immunotherapy strategies for TC cancer patients.

This review aims to provide insight into the role of N6-methyladenosine (m6A) modification in neoplastic immunity and subsequent tumorigenesis. m6A modification, which is catalyzed by methyltransferases, demethylases and reader proteins, has emerged as a widespread regulatory mechanism that controls immune-related gene expression and immune reactions during tumorigenesis. Aberrant m6A modification changes the neoplastic immune response in multiple cancers by regulating immune cell infiltration, tumor-promoting inflammation, immunosuppression, immune surveillance, and antitumor immune responses. m6A modification affects immune cell recruitment and cancer-promoting inflammation in hepatocellular carcinoma (HCC) to alter the progression of HCC. m6A modification has been implicated in the infiltration of immune cells and the activation of immune pathways, changing the proliferation and metastasis of gastric cancer. Immune surveillance and the antitumor immune response in breast cancer were enhanced via m6A modification, which inhibited tumor proliferation. m6A modification participates in neoplastic immunoregulation to influence tumor progression.

Comprehensive analysis of m6A regulator-based methylation modification patterns characterized by distinct immune profiles in colon adenocarcinomas

Cancer stemness, mainly consisting of chemo-resistance, radio-resistance, tumorigenesis, metastasis, tumor self-renewal, cancer metabolism reprogramming, and tumor immuno-microenvironment remodeling, play crucial roles in the cancer progression process and has become the hotspot of cancer research field in recent years. Nowadays, the exact molecular mechanisms of cancer stemness have not been fully understood. Extensive studies have recently implicated that non-coding RNA (ncRNA) plays vital roles in modulating cancer stemness. Notably, N6-methyladenosine (m6A) modification is of crucial importance for RNAs to exert their biological functions, including RNA splicing, stability, translation, degradation, and export. Emerging evidence has revealed that m6A modification can govern the expressions and functions of ncRNAs, consequently controlling cancer stemness properties. However, the interaction mechanisms between ncRNAs and m6A modification in cancer stemness modulation are rarely investigated. In this review, we elucidate the recent findings on the relationships of m6A modification, ncRNAs, and cancer stemness. We also focus on some key signaling pathways such as Wnt/β-catenin signaling, MAPK signaling, Hippo signaling, and JAK/STAT3 signaling to illustrate the underlying interplay mechanisms between m6A modification and ncRNAs in cancer stemness. In particular, we briefly highlight the clinical potential of ncRNAs and m6A modifiers as promising biomarkers and therapeutic targets for indicating cancer stemness properties and improving the diagnostic precision for a wide variety of cancers.

Multiple roles of p53 in cancer development: Regulation of tumor microenvironment, m6A modification and diverse cell death mechanisms

Recent studies have highlighted the biological significance of RNA N6-methyladenosine (m6A) modification in tumorigenicity and progression. However, it remains unclear whether m6A modifications also have potential roles in immune regulation and tumor microenvironment (TME) formation.Methods: In this study, we curated 23 m6A regulators and performed consensus molecular subtyping with NMF algorithm to determine m6A modification patterns and the m6A-related gene signature in colon cancer (CC). The ssGSEA and CIBERSORT algorithms were employed to quantify the relative infiltration levels of various immune cell subsets. An PCA algorithm based m6Sig scoring scheme was used to evaluate the m6A modification patterns of individual tumors with an immune response.Results: Three distinct m6A modification patterns were identified among 1307 CC samples, which were also associated with different clinical outcomes and biological pathways. The TME characterization revealed that the identified m6A patterns were highly consistent with three known immune profiles: immune-inflamed, immune-excluded, and immune-desert, respectively. Based on the m6Sig score, which was extracted from the m6A-related signature genes, CC patients can be divided into high and low score subgroups. Patients with lower m6Sig score was characterized by prolonged survival time and enhanced immune infiltration. Further analysis indicated that lower m6Sig score also correlated with greater tumor mutation loads, PD-L1 expression, and higher mutation rates in SMGs (e.g., PIK3CA and SMAD4). In addition, patients with lower m6Sig scores showed a better immune responses and durable clinical benefits in three independent immunotherapy cohorts.Conclusions: This study highlights that m6A modification is significantly associated with TME diversity and complexity. Quantitatively evaluating the m6A modification patterns of individual tumors will strengthen our understanding of TME characteristics and promote more effective immunotherapy strategies.

Random Forest model reveals the interaction between N6-methyladenosine modifications and RNA-binding proteins

The specific functions of RNA N6-methyladenosine (m6A) modifications in the glioma tumor microenvironment (TME) and glioma patient prognosis and treatment have not been determined to date. The objective of the study was to determine the role of m6A modifications in glioma TME. Nonnegative matrix factorization (NMF) methods were used to determine m6A clusters and m6A gene signatures based on 21 genes relating to m6A modifications. TME characteristics for each m6A cluster and m6A gene signature were quantified by established m6A score. The utility of m6A score was validated in immunotherapy and other antiangiogenic treatment cohorts. Three m6A clusters were identified among 3,395 glioma samples, and they were linked to different biological activities and clinical outcomes. The m6A clusters were highly consistent with immune profiles known as immune-inflamed, immune-excluded, and immune-desert phenotypes. Clusters within individual tumors could predict glioma inflammation, molecular subtypes, TME stromal activity, genetic variation, alternative splicing, and prognosis. As for the m6A score and m6A gene signature, patients with low m6A scores exhibited an increased tumor mutation burden, immune activity, neoantigen load, and prolonged survival. A low m6A score indicated the potential for a low level of T-cell dysfunction, a considerably better treatment response, and durable clinical benefits from immunotherapy, bevacizumab and regorafenib. Glioma m6A clusters and gene signatures have distinctive TME features. The m6A gene signature may guide prognostic assessments and promote the use of effective strategies.

BackgroundSeveral recent studies have confirmed epigenetic regulation of the immune response. However, the potential role of RNA N6-methyladenosine (m6A) modifications in cervical cancer and tumour microenvironment (TME) cell infiltration remain unclear.ResultsWe evaluated and analysed m6A modification patterns in 307 cervical cancer samples from The Cancer Genome Atlas (TCGA) dataset based on 13 m6A regulators. Pearson correlation analysis was used to identify lncRNAs associated with m6A, followed by univariate Cox regression analysis to screen their prognostic role in cervical cancer patients. We also correlated TME cell infiltration characteristics with modification patterns. We screened six m6A-associated lncRNAs as prognostic lncRNAs and established the prognostic profile of m6A-associated lncRNAs by least absolute shrinkage and choice of operator (LASSO) Cox regression. The corresponding risk scores of the patients were derived based on their prognostic features, and the correlation between this feature model and disease prognosis was analysed. The prognostic model constructed based on the TCGA-CESC (The Cancer Genome Cervical squamous cell carcinoma and endocervical adenocarcinoma) dataset showed strong prognostic power in the stratified analysis and was confirmed as an independent prognostic indicator for predicting the overall survival of patients with CESC. Enrichment analysis showed that biological processes, pathways, and markers associated with malignancy were more common in the high-risk subgroup. Risk scores were strongly correlated with the tumour grade. ECM receptor interactions and pathways in cancer were enriched in Cluster 2, while oxidative phosphorylation and other biological processes were enriched in Cluster 1. The expression of immune checkpoint molecules, including programmed death 1 (PD-1) and programmed death ligand 1 (PD-L1), was significantly increased in the high-risk subgroup, suggesting that this prognostic model could be a predictor of immunotherapy.ConclusionsThis study reveals that m6A modifications play an integral role in the diversity and complexity of TME formation. Assessing the m6A modification patterns of individual tumours will help improve our understanding of TME infiltration characteristics and thus guide immunotherapy more effectively. We also developed an independent prognostic model based on m6A-associated lncRNAs as a predictor of overall survival, which can also be used as a predictor of immunotherapy.

N6-methyladenosine-dependent upregulation of lnc-28509 increases apoptosis and suppresses bacterial infection in sea cucumber.

Stress-adaptive mechanisms enabling cancer cells to survive under glucose deprivation remain elusive. N6-methyladenosine (m6A) modification plays important roles in determining cancer cell fate and cellular stress response to nutrient deficiency. However, whether m6A modification functions in the regulation of cancer cell survival under glucose deprivation is unknown. Here, we found that glucose deprivation reduced m6A modification levels. Increasing m6A modification resulted in increased hepatoma cell necrosis under glucose deprivation, whereas decreasing m6A modification had an opposite effect. Integrated m6A-seq and RNA-seq revealed potential targets of m6A modification under glucose deprivation, including the transcription factor FOSL1; further, glucose deprivation upregulated FOSL1 by inhibiting FOSL1 mRNA decay in an m6A-YTHDF2-dependent manner through reducing m6A modification in its exon1 and 5'-UTR regions. Functionally, FOSL1 protected hepatoma cells against glucose deprivation-induced necrosis in vitro and in vivo. Mechanistically, FOSL1 transcriptionally repressed ATF3 by binding to its promoter. Meanwhile, ATF3 and MAFF interacted via their leucine zipper domains to form a heterodimer, which competed with NRF2 for binding to antioxidant response elements in the promoters of NRF2 target genes, thereby inhibiting their transcription. Consequently, FOSL1 reduced the formation of the ATF3-MAFF heterodimer, thereby enhancing NRF2 transcriptional activity and the antioxidant capacity of glucose-deprived-hepatoma cells. Thus, FOSL1 alleviated the necrosis-inducing effect of glucose deprivation-induced reactive oxygen species accumulation. Collectively, our study uncovers the protective role of m6A-FOSL1-ATF3 axis in hepatoma cell necrosis under glucose deprivation, and may provide new targets for cancer therapy.

N6-methyladenosine (m6A) modification is one of the most prevalent forms of epigenetic modification and plays an important role in the development of degenerative diseases such as osteoarthritis (OA). However, the evidence concerning the role of m6A modification in OA is insufficient. Here, m6A modification was increased in human OA cartilage and degenerated chondrocytes. Among all of the m6A enzymes, the expression of the demethylase fat mass and obesity-associated protein (FTO) decreased dramatically. Conditional knockout of FTO in chondrocytes accelerates OA progression. FTO transcription is regulated by runt-related transcription factor-1 (RUNX1). Reduced FTO elevates m6A modification at the adenosine N6 position in SMAD family member 2 (SMAD2) mRNA, whose stability is subsequently modulated by the recruited m6A reader protein YTH N6-methyladenosine RNA binding protein F2 (YTHDF2). Collectively, these findings reveal the function and mechanism of the m6A family member FTO in OA progression. Therefore, reducing m6A modification to increase SMAD2 stability by activating FTO might be a potential therapeutic strategy for OA treatment.

Diabetic foot ulcer (DFU) is a common and serious complication of diabetes mellitus (DM) and one of the main causes of disability or mortality in diabetic patients. Astragaloside IV (AS-IV) is a naturally occurring plant product that can be used to treat DM and its complications. N6-methyladenosine (m6A) modification participates in the progression of DM. This study aimed to explore the potential molecular mechanisms of m6A modification induced by AS-IV treatment of DFU. The biological behaviours of cells were assessed by cell counting kit-8, EdU incorporation, wound healing, and transwell assays. The levels of m6A modification and relative genes mRNA were detected by dot blot assay and quantitative real-time PCR. The interactions between molecules were evaluated by methylated RNA immunoprecipitation (MeRIP), RIP, fluorescence insitu hybridisation, and dual-luciferase reporter analysis. The results suggested AS-IV facilitated cell proliferation and migration of AGE-induced HaCaT cells and accelerated wound healing, thereby alleviating DFU progression. Mechanically, AS-IV promoted the m6A modification levels of SIRT1 invivo and invitro which was associated with increased METTL3. Besides, SIRT1 was the target of METTL3-mediated m6A modification. Silencing of SIRT1 abrogated the biological effects of METTL3. The mitigation effect of AS-IV on DFU is achieved by promoting m6A modification of SIRT1 to enhance SIRT1 stability. This study not only reveals the novel molecular mechanisms of AS-IV in the treatment of DFU but also provides new insights for the treatment of diabetes and its complications.

Nuclear retention coupled with sequential polyadenylation dictates post-transcriptional m6A modification in the nucleus