P53-associated lncRNAs regulate immune functions and RNA-modifiers in gastric cancer.

TP53, a guardian of the genome, suppresses or enhances tumors through various regulatory pathways. However, the role of p53-related long non-coding RNAs (lncRNAs) in immune regulation of tumor microenvironment and prognosis of gastric cancer (GC) is so far unelucidated. We analyzed the role of TP53-associated lncRNAs (obtained from the TP53LNC-DB database) in immune regulation, immune cell infiltration and RNA modification in gastric cancer. Firstly, using multivariate COX regression analysis, we identified eight lncRNAs related to the prognosis of GC. Furthermore, based on the expression of the lncRNA signature and risk score, the GC patients were divided into high-risk and low-risk groups. We found that M2-macrophages have significantly higher infiltration in the high-risk group. Similarly, significant differences in immune function (APC_co_stimulation, CCR, and checkpoint) and m6A modification (FTO, ZC3H13, YTHDC1, and RBM15), and m5C modification (NOP2 and TET1) between both groups were also observed. These signature lncRNAs were also positively associated with oxidative stress-related genes (MPO, MAPK14, HMOX1, and APP). Additionally, we found that high expression of GAS5 and low expression of MALAT1 in Helicobacter pylori (H-pylori) positive GC patients. Finally, GC patients in the low-risk group showed higher resistance to immunotherapy while patients in the high-risk group were more sensitive to various chemotherapy drugs. Based on these findings, we conclude that p53-associated lncRNAs signature could potentially predict the immune status and overall survival, and may also be used for risk management and planning immunotherapy for gastric cancer patients.

Implications of m5C modifications in ribosomal proteins on oxidative stress, metabolic reprogramming, and immune responses in patients with mid-to-late-stage head and neck squamous cell carcinoma: Insights from nanopore sequencing.

Background: Head and Neck Squamous Cell Carcinoma (HNSCC) is a malignancy characterized by a high incidence and recurrence rate. 5-methylcytosine (m5C) RNA modification is a common alteration affecting cancer progression; however, how m5C operates within the tumor microenvironment of HNSCC remains to be elucidated. Methods: We conducted Nanopore sequencing on 3 pairs of cancer and paracancerous tissues from mid- and late-stage HNSCC, obtaining 132 upregulated genes (transcriptomically upregulated, m5C elevated) and 129 downregulated genes (transcriptomically downregulated, m5C reduced). Subsequent Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were performed; a differential gene interaction network (PPI) was constructed, revealing the interactions of each gene with others in the network. Co-expression analysis was performed on the genes within the PPI, unveiling their expression and regulatory relationships. Through GSVA analysis, variations in related pathways under different states were identified. Furthermore, results of m5C in lncRNA were screened, followed by target gene prediction. Results: Sequencing results from the 3 pairs of mid- and late-stage HNSCC cancer and paracancerous tissues demonstrated that RPS27A, RPL8, and the lncRNAs including differentiation antagonizing nonprotein coding RNA (DANCR), DCST1 antisense RNA 1 (CCDC144NL-AS1), Growth Arrest-Specific Transcript 5 (GAS5), Nuclear Paraspeckle Assembly Transcript 1 (NEAT1), and Small Nucleolar RNA Host Gene 3 (SNHG3), etc., under m5Cregulation, have close connections with surrounding genes. The differentially m5Cmodified genes are primarily involved in ribosomal protein synthesis, oxidative stress response, metabolic reprogramming, immunity, and other life processes; pathways like mitochondrial protein import and photodynamic therapy induced unfolded protein response are upregulated in the tumor, while pathways, including the classic P53, are suppressed. Analysis on m5C-regulated long non-coding RNAs (lncRNAs) revealed tight associations with RPS27A and RPL8 as well. Conclusion: Our study identifies the key factors and signaling pathways involving m5C in HNSCC. The findings suggest that ribosome-related genes might regulate ribosomal protein synthesis, oxidative stress response, metabolic reprogramming, and immune response through m5C RNA modification by means like hypoxia and ferroptosis, thereby playing a pivotal role in the onset and progression of HNSCC. Hence, attention should be paid to the role of ribosomes in HNSCC. These findings may facilitate the precision and individualized treatment of patients with mid- and late-stage HNSCC in clinical settings.

m5c-iDeep: 5-Methylcytosine sites identification through deep learning.

5-Methylcytosine (m5c) is a modified cytosine base which is formed as the result of addition of methyl group added at position 5 of carbon. This modification is one of the most common PTM that used to occur in almost all types of RNA. The conventional laboratory methods do not provide quick reliable identification of m5c sites. However, the sequence data readiness has made it feasible to develop computationally intelligent models that optimize the identification process for accuracy and robustness. The present research focused on the development of in-silico methods built using deep learning models. The encoded data was then fed into deep learning models, which included gated recurrent unit (GRU), long short-term memory (LSTM), and bi-directional LSTM (Bi-LSTM). After that, the models were subjected to a rigorous evaluation process that included both independent set testing and 10-fold cross validation. The results revealed that LSTM-based model, m5c-iDeep, outperformed revealing 99.9 % accuracy while comparing with existing m5c predictors. In order to facilitate researchers, m5c-iDeep was also deployed on a web-based server which is accessible at https://taseersuleman-m5c-ideep-m5c-ideep.streamlit.app/.

m5C RNA methylation: a potential mechanism for infectious Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder caused by a variety of factors, including age, genetic susceptibility, cardiovascular disease, traumatic brain injury, and environmental factors. The pathogenesis of AD is largely associated with the overproduction and accumulation of amyloid-ß peptides and the hyperphosphorylation of tau protein in the brain. Recent studies have identified the presence of diverse pathogens, including viruses, bacteria, and parasites, in the tissues of AD patients, underscoring the critical role of central nervous system infections in inducing pathological changes associated with AD. Nevertheless, it remains unestablished about the specific mechanism by which infections lead to the occurrence of AD. As an important post-transcriptional RNA modification, RNA 5-methylcytosine (m5C) methylation regulates a wide range of biological processes, including RNA splicing, nuclear export, stability, and translation, therefore affecting cellular function. Moreover, it has been recently demonstrated that multiple pathogenic microbial infections are associated with the m5C methylation of the host. However, the role of m5C methylation in infectious AD is still uncertain. Therefore, this review discusses the mechanisms of pathogen-induced AD and summarizes research on the molecular mechanisms of m5C methylation in infectious AD, thereby providing new insight into exploring the mechanism underlying infectious AD.

NXPH4 mediated by m5C contributes to the malignant characteristics of colorectal cancer via inhibiting HIF1A degradation.

OBJECTIVE: Colorectal cancer (CRC) is a form of malignancy that exhibits a comparatively elevated occurrence and fatality rate. Given the relatively slower progress in diagnostic and therapeutic approaches for CRC, there is a need to investigate more accurate and efficient biomarkers. METHODS: Core regulatory genes were screened using the TCGA database, and the expression of neurexophilin 4 (NXPH4) and its prognostic implications were validated using tissue microarray staining. The assessment of NXPH4 functions involved a range of experiments, including cellular, organoid, and murine models. Furthermore, a regulatory network between m5C, NXPH4, and HIF1A was established through several in vitro experiments. RESULTS: The overexpression of NXPH4 is associated with unfavorable prognoses in patients with CRC and hepatocellular carcinoma. Additionally, it facilitates the progression of malignant tumors both in laboratory settings and in living organisms of colorectal carcinoma. Our research also reveals that NXPH4 mRNA can avoid degradation through RNautophagy, relying on an m5C-dependent mechanism. Moreover, NXPH4 amplifies the HIF signaling pathway and stabilizes HIF1A by competitively binding to PHD4. CONCLUSIONS: NXPH4, regulated by m5C, promotes malignant tumor progression and regulates the HIF pathway. Consequently, targeting NXPH4 through molecular therapies could potentially serve as an efficacious therapeutic strategy for the management of CRC exhibiting elevated NXPH4 expression.

A novel m6A/m5C/m1A/m7G-related classification and risk signature predicts prognosis and reveals immunotherapy inclination in gastric cancer.

Background: Gastric cancer (GC) is characterized by high morbidity and mortality rates, and the prognosis is not optimistic. Therefore, the search for new biomarkers is crucial. Methylation modifications in RNA modifications play a crucial role in tumors. However, the role of methylation modification of integrated m6A/m5C/m1A/m7G, in GC and its related analysis have not been reported. It still needs to be studied in depth. Our study aims to deepen our understanding of m6A/m5C/m1A/m7G methylation and potentially provide new strategies for GC treatment. Methods: We used TCGA-STAD (The Cancer Genome Atlas-Stomach Adenocarcinoma) as a training set and GSE84433 as a validation set to analyze and determine potential associations between m6A/m5C/m1A/m7G-related genes and clinical risk of GC. In addition, we explored the prognostic value and potential biological mechanisms of m6A/m5C/m1A/m7G-related genes in GC through consistent clustering, differential expression gene identification, enrichment analysis, and immune infiltration analysis. Finally, we constructed m6A/m5C/m1A/m7G-related risk signature (MRRS) to evaluate the correlation between risk grade and survival prognosis, drug sensitivity, and immune infiltration, and validated the validity by immunohistochemical staining. Results: We identified subgroups of C1, C2, and C3 patients by consensus clustering using data from 45 m6A/m5C/m1A/m7G-related genes. The three groups showed significant differences in survival, immune scores, and immune cell infiltration. We then constructed MRRS using least absolute shrinkage and selection operator (LASSO) regression analysis, including SLC5A6, FKBP10, GPC3, and GGH, which could accurately differentiate between high-/low-risk populations. Its accuracy was further validated in the validation set and immunohistochemical staining. These results suggest that m6A/m5C/m1A/m7G are closely related to the GC tumor immune microenvironment, and MRRS has good performance in predicting the survival of GC patients. Conclusions: In this study, we highlighted the association of m6A/m5C/m1A/m7G subtypes with changes in the GC immunotumor microenvironment. We constructed and validated MRRS, which is valuable in predicting survival, immune infiltration and drug sensitivity in GC patients. This helps to deepen our understanding of m6A/m5C/m1A/m7G methylation and potentially provides new strategies for GC treatment.

Prognostic signature and immune efficacy of m1A-, m5C-, m6A-, m7G-, and DNA methylation-related regulators in hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) is the main type of primary liver cancer, and its related death ranks third worldwide. The curative methods and progress prediction markers of HCC are not sufficient enough. Nevertheless, little progress has been made in the signature of m1A-, m5C-, m6A-, m7G-, and DNA methylation of HCC. Results: We calibrated a risk gene signature model that can be used to categorize HCC patients based on univariate, multivariate, and LASSO Cox regression analysis. This gene signature classified the patients into high- and low-risk subgroups. Patients in the high-risk group showed significantly reduced overall survival (OS) compared with patients in the low-risk group. The gene set variation analysis (GSVA), immune infiltration, and immunotherapy response were analyzed. The results demonstrated that an immunosuppressive environment was exited and the high-risk group had higher sensitivity to 5-fluorouracil, cisplatin, sorafenib, tamoxifen, and epirubicin. These results indicated personalized therapy should be taken into consideration. Conclusions: Our findings enriched our understanding of the molecular heterogeneity, tumor microenvironment (TME), and drug susceptibility of HCC. m1A-, m5C-, m6A-, m7G-, and DNA methylation-related regulators may be promising biomarkers for future research.

Base-resolution m5C profiling across the mammalian transcriptome by bisulfite-free enzyme-assisted chemical labeling approach.

5-methylcytosine (m5C) is a prevalent RNA modification crucial for gene expression regulation. However, accurate and sensitive m5C sites identification remains challenging due to severe RNA degradation and reduced sequence complexity during bisulfite sequencing (BS-seq). Here, we report m5C-TAC-seq, a bisulfite-free approach combining TET-assisted m5C-to-f5C oxidation with selective chemical labeling, therefore enabling direct base-resolution m5C detection through pre-enrichment and C-to-T transitions at m5C sites. With m5C-TAC-seq, we comprehensively profiled the m5C methylomes in human and mouse cells, identifying a substantially larger number of confident m5C sites. Through perturbing potential m5C methyltransferases, we deciphered the responsible enzymes for most m5C sites, including the characterization of NSUN5's involvement in mRNA m5C deposition. Additionally, we characterized m5C dynamics during mESC differentiation. Notably, the mild reaction conditions and preservation of nucleotide composition in m5C-TAC-seq allow m5C detection in chromatin-associated RNAs. The accurate and robust m5C-TAC-seq will advance research into m5C methylation functional investigation.

Autophagy Deficiency Induced by SAT1 Potentiates Tumor Progression in Triple-Negative Breast Cancer.

Aggressive triple-negative breast cancer (TNBC) still lacks approved targeted therapies, requiring more exploration of its underlying mechanisms. Previous studies have suggested a potential role of SAT1 (Spermidine/Spermine N1-acetyltransferase 1) in cancer, which needs to be further elucidated in breast cancer. In this study, highly expressed SAT1 in TNBC signified worse patient prognoses. And SAT1 knockdown effectively inhibited the proliferation and migration abilities of TNBC cells in vitro and in vivo. In terms of mechanism, the transcription factor JUN enhanced SAT1 transcriptional activity by binding to its promoter region. Then, SAT1 protein in the cytoplasm engaged in directly binding with YBX1 for sustaining YBX1 protein stability via deubiquitylation mediated by the E3 ligase HERC5. Further, SAT1 was found to suppress autophagy remarkably via stabilization of mTOR mRNA with the accumulation of YBX1-mediated methyl-5-cytosine (m5C) modification. These findings proved that SAT1 drives TNBC progression through the SAT1/YBX1/mTOR axis, which may provide a potential candidate for targeted therapy in advanced TNBC.

m6A- and m5C- modified lncRNAs orchestrate the prognosis in cutaneous melanoma and m6A- modified LINC00893 regulates cutaneous melanoma cell metastasis.

BACKGROUND: As the most important modifications on the RNA level, N6-methyladenosine (m6A-) and 5-methylcytosine (m5C-) modification could have a direct influence on the RNAs. Long non-coding RNAs (lncRNAs) could also be modified by methylcytosine modification. Compared with mRNAs, the function of lncRNAs could be more potent to some extent in biological processes like tumorigenesis. Until now, rare reports have been done associated with cutaneous melanoma. Herein, we wonder if the m6A- and m5C- modified lncRNAs could influence the immune landscape and prognosis in melanoma, and we also want to find some lncRNAs which could directly affect the malignant behaviors of melanoma. METHODS: Systematically, we explored the expression pattern of m6A- and m5C- modified lncRNAs in melanoma from datasets including UCSC Xena and NCBI GEO, and the prognostic lncRNAs were selected. Then, according to the expression pattern of lncRNAs, melanoma samples from these datasets were divided into several subtypes. Prognostic model, nomogram survival model, drug sensitivity, GO, and KEGG pathway analysis were performed. Furthermore, among several selected lncRNAs, we identified one lncRNA named LINC00893 and investigated its expression pattern and its biological function in melanoma cell lines. RESULTS: We identified 27 m6A- and m5C- related lncRNAs which were significantly associated with survival, and we made a subtype analysis of melanoma samples based on these 27 lncRNAs. Among the two subtypes, we found differences of immune cells infiltration between these two subtypes. Then, LASSO algorithm was used to screen the optimized lncRNAs combination including ZNF252P-AS1, MIAT, FAM13A-AS1, LINC-PINT, LINC00893, AGAP2-AS1, OIP5-AS1, and SEMA6A-AS1. We also found that there was a significant correlation between the different risk groups predicted based on RS model and the actual prognosis. The nomogram survival model based on independent survival prognostic factors was also constructed. Besides, sensitivity to chemotherapeutic agents, GO and KEGG analysis were performed. In different risk groups, a total of 14 drug molecules with different distributions were obtained, which included AZD6482, AZD7762, AZD8055, camptothecin, dasatinib, erlotinib, gefitinib, gemcitabine, GSK269962A, nilotinib, rapamycin, and sorafenib. A total of 55 significantly related biological processes and 17 KEGG signaling pathways were screened. At last, we noticed that LINC00893 had a relatively lower expression in melanoma tissue and cell lines compared with adjacent tissues and epidermal melanocyte, and down-regulation of LINC00893 could promote the malignant behavior of melanoma cells in A875 and MV3. In these two melanoma cell lines, down-regulation of m6A-related molecules like YTHDF3 and METTL3 could promote the expression of LINC00893. CONCLUSION: We made an analysis of m6A- and m5C- related lncRNAs in melanoma samples and a prediction of these lncRNAs' role in prognosis, tumor microenvironment, immune infiltration, and clinicopathological features. We also found that LINC00893, which is potentially regulated by m6A modification, could serve as a tumor-suppressor in melanoma and play an inhibitory role in melanoma metastasis.

NSUN6-mediated 5-methylcytosine modification of NDRG1 mRNA promotes radioresistance in cervical cancer.

BACKGROUND: Radioresistance is the leading cause of death in advanced cervical cancer (CC). Dysregulation of RNA modification has recently emerged as a regulatory mechanism in radiation and drug resistance. We aimed to explore the biological function and clinical significance of 5-methylcytosine (m5C) in cervical cancer radiosensitivity. METHODS: The abundance of RNA modification in radiotherapy-resistant and sensitive CC specimens was quantified by liquid chromatography-tandem mass spectrometry. The essential RNA modification-related genes involved in CC radiosensitivity were screened via RNA sequencing. The effect of NSUN6 on radiosensitivity was verified in CC cell lines, cell-derived xenograft (CDX), and 3D bioprinted patient-derived organoid (PDO). The mechanisms of NSUN6 in regulating CC radiosensitivity were investigated by integrative m5C sequencing, mRNA sequencing, and RNA immunoprecipitation. RESULTS: We found a higher abundance of m5C modification in resistant CC samples, and NSUN6 was the essential m5C-regulating gene concerning radiosensitivity. NSUN6 overexpression was clinically correlated with radioresistance and poor prognosis in cervical cancer. Functionally, higher NSUN6 expression was associated with radioresistance in the 3D PDO model of cervical cancer. Moreover, silencing NSUN6 increased CC radiosensitivity in vivo and in vitro. Mechanistically, NDRG1 was one of the downstream target genes of NSUN6 identified by integrated m5C-seq, mRNA-seq, and functional validation. NSUN6 promoted the m5C modification of NDRG1 mRNA, and the m5C reader ALYREF bound explicitly to the m5C-labeled NDRG1 mRNA and enhanced NDRG1 mRNA stability. NDRG1 overexpression promoted homologous recombination-mediated DNA repair, which in turn led to radioresistance in cervical cancer. CONCLUSIONS: Aberrant m5C hypermethylation and NSUN6 overexpression drive resistance to radiotherapy in cervical cancer. Elevated NSUN6 expression promotes radioresistance in cervical cancer by activating the NSUN6/ALYREF-m5C-NDRG1 pathway. The low expression of NSUN6 in cervical cancer indicates sensitivity to radiotherapy and a better prognosis.

Dynamic RNA methylation modifications and their regulatory role in mammalian development and diseases.

Among over 170 different types of chemical modifications on RNA nucleobases identified so far, RNA methylation is the major type of epitranscriptomic modifications existing on almost all types of RNAs, and has been demonstrated to participate in the entire process of RNA metabolism, including transcription, pre-mRNA alternative splicing and maturation, mRNA nucleus export, mRNA degradation and stabilization, mRNA translation. Attributing to the development of high-throughput detection technologies and the identification of both dynamic regulators and recognition proteins, mechanisms of RNA methylation modification in regulating the normal development of the organism as well as various disease occurrence and developmental abnormalities upon RNA methylation dysregulation have become increasingly clear. Here, we particularly focus on three types of RNA methylations: N6-methylcytosine (m6A), 5-methylcytosine (m5C), and N7-methyladenosine (m7G). We summarize the elements related to their dynamic installment and removal, specific binding proteins, and the development of high-throughput detection technologies. Then, for a comprehensive understanding of their biological significance, we also overview the latest knowledge on the underlying mechanisms and key roles of these three mRNA methylation modifications in gametogenesis, embryonic development, immune system development, as well as disease and tumor progression.

Epitranscriptomic cytidine methylation of the hepatitis B viral RNA is essential for viral reverse transcription and particle production.

Epitranscriptomic RNA modifications have emerged as important regulators of the fate and function of viral RNAs. One prominent modification, the cytidine methylation 5-methylcytidine (m5C), is found on the RNA of HIV-1, where m5C enhances the translation of HIV-1 RNA. However, whether m5C functionally enhances the RNA of other pathogenic viruses remains elusive. Here, we surveyed a panel of commonly found RNA modifications on the RNA of hepatitis B virus (HBV) and found that HBV RNA is enriched with m5C as well as ten other modifications, at stoichiometries much higher than host messenger RNA (mRNA). Intriguingly, m5C is mostly found on the epsilon hairpin, an RNA element required for viral RNA encapsidation and reverse transcription, with these m5C mainly deposited by the cellular methyltransferase NSUN2. Loss of m5C from HBV RNA due to NSUN2 depletion resulted in a partial decrease in viral core protein (HBc) production, accompanied by a near-complete loss of the reverse transcribed viral DNA. Similarly, mutations introduced to remove the methylated cytidines resulted in a loss of HBc production and reverse transcription. Furthermore, pharmacological disruption of m5C deposition led to a significant decrease in HBV replication. Thus, our data indicate m5C methylations as a critical mediator of the epsilon elements' function in HBV virion production and reverse transcription, suggesting the therapeutic potential of targeting the m5C methyltransfer process on HBV epsilon as an antiviral strategy.

NSUN4 mediated RNA 5-methylcytosine promotes the malignant progression of glioma through improving the CDC42 mRNA stabilization.

5-Methylcytosine (m5C) methylation is a significant post-transcriptional modification that play a crucial role in the development and progression of numerous cancers. Whereas the functions and molecular mechanisms underlying m5C methylation in gliomas remain unclear. This study dedicated to explore changes of m5C levels and the clinical significance of the m5C writer NSUN4 in gliomas. We found that high m5C levels were negatively related to prognosis of patients with glioma. Moreover, gain- and loss-of-function experiments revealed the role of NSUN4 in enhancing m5C modification of mRNA to promote the malignant progression of glioma. Mechanistically speaking, NSUN4-mediated m5C alterations regulated ALYREF binding to CDC42 mRNA, thereby impacting the mRNA stability of CDC42. We also demonstrated that CDC42 promoted glioma proliferation, migration, and invasion by activating the PI3K-AKT pathway. Additionally, rescue experiments proved that CDC42 overexpression weaken the inhibitory effect of NSUN4 knockdown on the malignant progression of gliomas in vitro and in vivo. Our findings elucidated that NSUN4-mediated high m5C levels promote ALYREF binding to CDC42 mRNA and regulate its stability, thereby driving the malignant progression of glioma. This provides theoretical support for targeted the treatment of gliomas.

YBX1 promotes homologous recombination and resistance to platinum-induced stress in ovarian cancer by recognizing m5C modification.

Platinum-based chemotherapy causes genetic damage and induces apoptosis in ovarian cancer cells. Enhancing the ability to resist platinum drug-induced DNA damage and apoptotic stress is critical for tumor cells to acquire drug resistance. Here, we found that Y-box binding protein 1 (YBX1) was highly expressed in cisplatin-resistant patient-derived organoids (PDOs) and was a crucial gene for alleviating platinum-induced stress and maintaining drug resistance characteristics in ovarian cancer cells. Mechanistically, YBX1 recognized m5C modifications in CHD3 mRNA and maintained mRNA stability by recruiting PABPC1 protein. This regulatory process enhanced chromatin accessibility and improved the efficiency of homologous recombination (HR) repair, facilitating tumor cells to withstand platinum-induced apoptotic stress. In addition, SU056, an inhibitor of YBX1, exhibited the potential to reverse platinum resistance in subcutaneous and PDO orthotopic xenograft models. In conclusion, YBX1 is critical for ovarian cancer cells to alleviate the platinum-induced stress and may be a potential target for reversing drug-resistant therapies.

RNA m5C methylation: a potential modulator of innate immune pathways in hepatocellular carcinoma.

RNA 5-methylcytosine (m5C) methylation plays a crucial role in hepatocellular carcinoma (HCC). As reported, aberrant m5C methylation is closely associated with the progression, therapeutic efficacy, and prognosis of HCC. The innate immune system functions as the primary defense mechanism in the body against pathogenic infections and tumors since it can activate innate immune pathways through pattern recognition receptors to exert anti-infection and anti-tumor effects. Recently, m5C methylation has been demonstrated to affect the activation of innate immune pathways including TLR, cGAS-STING, and RIG-I pathways by modulating RNA function, unveiling new mechanisms underlying the regulation of innate immune responses by tumor cells. However, research on m5C methylation and its interplay with innate immune pathways is still in its infancy. Therefore, this review details the biological significance of RNA m5C methylation in HCC and discusses its potential regulatory relationship with TLR, cGAS-STING, and RIG-I pathways, thereby providing fresh insights into the role of RNA methylation in the innate immune mechanisms and treatment of HCC.

RNA m5C methylation modification: a potential therapeutic target for SARS-CoV-2-associated myocarditis.

The Corona Virus Disease (COVID-19), caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), has quickly spread worldwide and resulted in significant morbidity and mortality. Although most infections are mild, some patients can also develop severe and fatal myocarditis. In eukaryotic RNAs, 5-methylcytosine (m5C) is a common kind of post-transcriptional modification, which is involved in regulating various biological processes (such as RNA export, translation, and stability maintenance). With the rapid development of m5C modification detection technology, studies related to viral m5C modification are ever-increasing. These studies have revealed that m5C modification plays an important role in various stages of viral replication, including transcription and translation. According to recent studies, m5C methylation modification can regulate SARS-CoV-2 infection by modulating innate immune signaling pathways. However, the specific role of m5C modification in SARS-CoV-2-induced myocarditis remains unclear. Therefore, this review aims to provide insights into the molecular mechanisms of m5C methylation in SARS-CoV-2 infection. Moreover, the regulatory role of NSUN2 in viral infection and host innate immune response was also highlighted. This review may provide new directions for developing therapeutic strategies for SARS-CoV-2-associated myocarditis.

NSUN2 affects diabetic retinopathy progression by regulating MUC1 expression through RNA m5C methylation.

BACKGROUND: Diabetic retinopathy (DR) is the leading cause of blinding eye disease among working adults and is primarily attributed to the excessive proliferation of microvessels, which leads to vitreous hemorrhage and retinal traction, thereby significantly impairing patient vision. NSUN2-mediated RNA m5C methylation is implicated in various diseases, and in this investigation, we focused on elucidating the impact of NSUN2 on the regulation of the expression of the downstream gene MUC1, specifically through RNA m5C methylation, on the progression of DR. METHOD: Utilizing Microarray analysis, we examined patient vitreous fluid to pinpoint potential therapeutic targets for DR. Differential expression of NSUN2 was validated through qRT-PCR, Western blot, and immunofluorescence in human tissue, animal tissue, and cell model of DR. The relationship between NSUN2 and DR was explored in vitro and in vivo through gene knockdown and overexpression. Various techniques, such as MeRIP-qPCR and dot blot, were applied to reveal the downstream targets and mechanism of action of NSUN2. RESULTS: The levels of both NSUN2 and RNA m5C methylation were significantly elevated in the DR model. Knockdown of NSUN2 mitigated DR lesion formation both in vitro and in vivo. Mechanistically, NSUN2 promoted MUC1 expression by binding to the RNA m5C reader ALYREF. Knockdown of ALYREF resulted in DR lesion alterations similar to those observed with NSUN2 knockdown. Moreover, MUC1 overexpression successfully reversed a series of DR alterations induced by NSUN2 silencing. CONCLUSIONS: NSUN2 regulates the expression of MUC1 through ALYREF-mediated RNA m5C methylation, thereby regulating the progression of DR and providing a new option for the treatment of DR in the future.

TET1 mediated m5C modification of RelB aggravates cerebral ischemia/reperfusion-induced neuroinflammation through regulating microglia polarization.

Microglia mediated neuroinflammation is one of the major contributors to brain damage in cerebral ischemia reperfusion injury (CI/RI). Recently, RNA modification was found to contribute to the regulation of microglia polarization and the subsequent development of cerebral I/R neuroinflammation. Herein, we investigated the effect and mechanism of m5C RNA modification in the microglia induced CI/RI neuroinflammation. We found that the m5C RNA modification levels decreased in the primary microglia isolated from a mouse model of intraluminal middle cerebral artery occlusion/reperfusion (MCAO/R) and the BV2 microglial cells subjected to oxygen-glucose deprivation and reoxygenation (OGD/R), and this change was accompanied by an increase in the M1/M2 polarization ratio. Furthermore, the expression of m5C demethylase TET1 in microglia increased, which promoted M1 polarization but impeded M2 polarization. Mechanistically, the higher TET1 expression decreased the m5C modification level of RelB and enhanced its mRNA stability, which subsequently increased the M1/M2 polarization ratio. In conclusion, this study provides insight into the role of m5C RNA modification in the pathogenesis of cerebral I/R neuroinflammation and may deepen our understanding on clinical therapy targeting the TET1-RelB axis.

Epitranscriptomic orchestrations: Unveiling the regulatory paradigm of m6A, A-to-I editing, and m5C in breast cancer via long noncoding RNAs and microRNAs.

Breast cancer (BC) poses a persistent global health challenge, particularly in countries with elevated human development indices linked to factors such as increased life expectancy, education, and wealth. Despite therapeutic progress, challenges persist, and the role of epitranscriptomic RNA modifications in BC remains inadequately understood. The epitranscriptome, comprising diverse posttranscriptional modifications on RNA molecules, holds the potential to intricately modulate RNA function and regulation, implicating dysregulation in various diseases, including BC. Noncoding RNAs (ncRNAs), acting as posttranscriptional regulators, influence physiological and pathological processes, including cancer. RNA modifications in long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) add an extra layer to gene expression control. This review delves into recent insights into epitranscriptomic RNA modifications, such as N-6-methyladenosine (m6A), adenine-to-inosine (A-to-I) editing, and 5-methylcytosine (m5C), specifically in the context of lncRNA and miRNAs in BC, highlighting their potential implications in BC development and progression. Understanding this intricate regulatory landscape is vital for deciphering the molecular mechanisms underlying BC and identifying potential therapeutic targets.