RESUMO
N6-methyladenosine (m6A) is a crucial RNA modification that regulates diverse biological processes in human cells, but its co-transcriptional deposition and functions remain poorly understood. Here, we identified the RNA helicase DDX21 with a previously unrecognized role in directing m6A modification on nascent RNA for co-transcriptional regulation. DDX21 interacts with METTL3 for co-recruitment to chromatin through its recognition of R-loops, which can be formed co-transcriptionally as nascent transcripts hybridize onto the template DNA strand. Moreover, DDX21's helicase activity is needed for METTL3-mediated m6A deposition onto nascent RNA following recruitment. At transcription termination regions, this nexus of actions promotes XRN2-mediated termination of RNAPII transcription. Disruption of any of these steps, including the loss of DDX21, METTL3, or their enzymatic activities, leads to defective termination that can induce DNA damage. Therefore, we propose that the R-loop-DDX21-METTL3 nexus forges the missing link for co-transcriptional modification of m6A, coordinating transcription termination and genome stability.
Assuntos
Adenosina , Adenosina/análogos & derivados , RNA Helicases DEAD-box , Exorribonucleases , Instabilidade Genômica , Metiltransferases , Estruturas R-Loop , RNA Polimerase II , Terminação da Transcrição Genética , Humanos , RNA Helicases DEAD-box/metabolismo , RNA Helicases DEAD-box/genética , Metiltransferases/metabolismo , Metiltransferases/genética , Adenosina/metabolismo , Adenosina/genética , Exorribonucleases/metabolismo , Exorribonucleases/genética , RNA Polimerase II/metabolismo , RNA Polimerase II/genética , Células HEK293 , Cromatina/metabolismo , Cromatina/genética , Dano ao DNA , Células HeLa , RNA/metabolismo , RNA/genética , Transcrição Gênica , Metilação de RNARESUMO
The mechanistic understanding of nascent RNAs in transcriptional control remains limited. Here, by a high sensitivity method methylation-inscribed nascent transcripts sequencing (MINT-seq), we characterized the landscapes of N6-methyladenosine (m6A) on nascent RNAs. We uncover heavy but selective m6A deposition on nascent RNAs produced by transcription regulatory elements, including promoter upstream antisense RNAs and enhancer RNAs (eRNAs), which positively correlates with their length, inclusion of m6A motif, and RNA abundances. m6A-eRNAs mark highly active enhancers, where they recruit nuclear m6A reader YTHDC1 to phase separate into liquid-like condensates, in a manner dependent on its C terminus intrinsically disordered region and arginine residues. The m6A-eRNA/YTHDC1 condensate co-mixes with and facilitates the formation of BRD4 coactivator condensate. Consequently, YTHDC1 depletion diminished BRD4 condensate and its recruitment to enhancers, resulting in inhibited enhancer and gene activation. We propose that chemical modifications of eRNAs together with reader proteins play broad roles in enhancer activation and gene transcriptional control.
Assuntos
Adenosina/análogos & derivados , Proteínas de Ciclo Celular/genética , Proteínas do Tecido Nervoso/genética , Fatores de Processamento de RNA/genética , RNA/genética , Fatores de Transcrição/genética , Adenosina/genética , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica/genética , Humanos , Metilação , Elementos Reguladores de Transcrição/genética , Ativação Transcricional/genéticaRESUMO
YTHDF proteins are main cytoplasmic 'reader' proteins of RNA N6-methyladenosine (m6A) methylation in mammals. They are largely responsible for m6A-mediated regulation in the cell cytosol by controlling both mRNA translation and degradation. Recent functional and mechanistic investigations of the YTHDF proteins revealed that these proteins have different functions to enable versatile regulation of the epitranscriptome. Their divergent functions largely originate from their different amino acid sequences in the low-complexity N termini. Consequently, they have different phase separation propensities and possess distinct post-translational modifications (PTMs). Different PTMs, subcellular localizations, and competition among partner proteins have emerged as three major mechanisms that control the functions of these YTHDF proteins. We also summarize recent progress on critical roles of these YTHDF proteins in anticancer immunity and the potential for targeting these proteins for developing new anticancer therapies.
Assuntos
Adenosina , Proteínas de Ligação a RNA , Humanos , Animais , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Adenosina/metabolismo , Adenosina/análogos & derivados , Processamento de Proteína Pós-Traducional , RNA/metabolismo , Metilação , Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/patologiaRESUMO
Modification of mRNA by N6-adenosine methylation (m6A) on internal bases influences gene expression in eukaryotes. How the dynamic genome-wide landscape of m6A-modified mRNAs impacts virus infection and host immune responses remains poorly understood. Here, we show that type I interferon (IFN) production triggered by dsDNA or human cytomegalovirus (HCMV) is controlled by the cellular m6A methyltrasferase subunit METTL14 and ALKBH5 demethylase. While METTL14 depletion reduced virus reproduction and stimulated dsDNA- or HCMV-induced IFNB1 mRNA accumulation, ALKBH5 depletion had the opposite effect. Depleting METTL14 increased both nascent IFNB1 mRNA production and stability in response to dsDNA. In contrast, ALKBH5 depletion reduced nascent IFNB1 mRNA production without detectably influencing IFN1B mRNA decay. Genome-wide transcriptome profiling following ALKBH5 depletion identified differentially expressed genes regulating antiviral immune responses, while METTL14 depletion altered pathways impacting metabolic reprogramming, stress responses, and aging. Finally, we determined that IFNB1 mRNA was m6A-modified within both the coding sequence and the 3' untranslated region (UTR). This establishes that the host m6A modification machinery controls IFNß production triggered by HCMV or dsDNA. Moreover, it demonstrates that responses to nonmicrobial dsDNA in uninfected cells, which shape host immunity and contribute to autoimmune disease, are regulated by enzymes controlling m6A epitranscriptomic changes.
Assuntos
DNA/imunologia , Regulação da Expressão Gênica/genética , Sistema Imunitário/enzimologia , Imunidade Inata/genética , Interferon beta/genética , Metiltransferases/metabolismo , Homólogo AlkB 5 da RNA Desmetilase/genética , Homólogo AlkB 5 da RNA Desmetilase/metabolismo , Animais , Linhagem Celular , Chlorocebus aethiops , Citomegalovirus/imunologia , Perfilação da Expressão Gênica , Humanos , Interferon beta/metabolismo , Estabilidade de RNA/genética , Células Vero , Replicação Viral/genéticaRESUMO
The N6-methyladenosine (m6 A) RNA modification serves crucial functions in RNA metabolism; however, the molecular mechanisms underlying the regulation of m6 A are not well understood. Here, we establish arginine methylation of METTL14, a component of the m6 A methyltransferase complex, as a novel pathway that controls m6 A deposition in mammalian cells. Specifically, protein arginine methyltransferase 1 (PRMT1) interacts with, and methylates the intrinsically disordered C terminus of METTL14, which promotes its interaction with RNA substrates, enhances its RNA methylation activity, and is crucial for its interaction with RNA polymerase II (RNAPII). Mouse embryonic stem cells (mESCs) expressing arginine methylation-deficient METTL14 exhibit significantly reduced global m6 A levels. Transcriptome-wide m6 A analysis identified 1,701 METTL14 arginine methylation-dependent m6 A sites located in 1,290 genes involved in various cellular processes, including stem cell maintenance and DNA repair. These arginine methylation-dependent m6 A sites are associated with enhanced translation of genes essential for the repair of DNA interstrand crosslinks; thus, METTL14 arginine methylation-deficient mESCs are hypersensitive to DNA crosslinking agents. Collectively, these findings reveal important aspects of m6 A regulation and new functions of arginine methylation in RNA metabolism.
Assuntos
Adenosina/análogos & derivados , Arginina/química , Metiltransferases/metabolismo , Células-Tronco Embrionárias Murinas/metabolismo , Processamento de Proteína Pós-Traducional , Proteína-Arginina N-Metiltransferases/metabolismo , RNA Polimerase II/metabolismo , Adenosina/química , Animais , Citoplasma , Metiltransferases/química , Metiltransferases/genética , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Proteína-Arginina N-Metiltransferases/genética , RNA Polimerase II/genética , TranscriptomaRESUMO
Papillary thyroid carcinoma (PTC) stands as the leading cancer type among endocrine malignancies, and there exists a strong correlation between thyroid cancer and obesity. However, the clinical significance and molecular mechanism of lipid metabolism in the development of PTC remain unclear. In this study, it was demonstrated that the downregulation of METTL16 enhanced lipid metabolism and promoted the malignant progression of PTC. METTL16 was expressed at lower levels in PTC tissues because of DNMT1-mediated hypermethylation of its promoter. Loss- and gain-of-function studies clarified the effects of METTL16 on PTC progression. METTL16 overexpression increased the abundance of m6A in SCD1 cells, increasing RNA decay via the m6A reader YTHDC2. The SCD1 inhibitor A939572 inhibited growth and slowed down lipid metabolism in PTC cells. These results confirm the crucial role of METTL16 in restraining PTC progression through SCD1-activated lipid metabolism in cooperation with YTHDC2. This suggests that the combination of METTL16 and anti-SCD1 blockade might constitute an effective therapy for PTC.
Assuntos
Metabolismo dos Lipídeos , Neoplasias da Glândula Tireoide , Humanos , Câncer Papilífero da Tireoide/genética , Câncer Papilífero da Tireoide/metabolismo , Metabolismo dos Lipídeos/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologia , Metilação de DNA , Linhagem Celular Tumoral , Proliferação de Células , Estearoil-CoA Dessaturase/genética , Estearoil-CoA Dessaturase/metabolismo , RNA Helicases/genética , RNA Helicases/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismoRESUMO
γδ T cells are an abundant T cell population at the mucosa and are important in providing immune surveillance as well as maintaining tissue homeostasis. However, despite γδ T cells' origin in the thymus, detailed mechanisms regulating γδ T cell development remain poorly understood. N6-methyladenosine (m6A) represents one of the most common posttranscriptional modifications of messenger RNA (mRNA) in mammalian cells, but whether it plays a role in γδ T cell biology is still unclear. Here, we show that depletion of the m6A demethylase ALKBH5 in lymphocytes specifically induces an expansion of γδ T cells, which confers enhanced protection against gastrointestinal Salmonella typhimurium infection. Mechanistically, loss of ALKBH5 favors the development of γδ T cell precursors by increasing the abundance of m6A RNA modification in thymocytes, which further reduces the expression of several target genes including Notch signaling components Jagged1 and Notch2. As a result, impairment of Jagged1/Notch2 signaling contributes to enhanced proliferation and differentiation of γδ T cell precursors, leading to an expanded mature γδ T cell repertoire. Taken together, our results indicate a checkpoint role of ALKBH5 and m6A modification in the regulation of γδ T cell early development.
Assuntos
Homólogo AlkB 5 da RNA Desmetilase , Linfócitos Intraepiteliais , RNA Mensageiro , Homólogo AlkB 5 da RNA Desmetilase/genética , Homólogo AlkB 5 da RNA Desmetilase/metabolismo , Animais , Linfócitos Intraepiteliais/enzimologia , Linfócitos Intraepiteliais/imunologia , Proteína Jagged-1/metabolismo , Camundongos , Camundongos Knockout , RNA Mensageiro/metabolismo , Receptor Notch2/metabolismo , Transdução de Sinais/genéticaRESUMO
The RNA modification N6-methyladenosine (m6A) regulates the interaction between RNA and various RNA binding proteins within the nucleus and other subcellular compartments and has recently been shown to be involved in experience-dependent plasticity, learning, and memory. Using m6A RNA-sequencing, we have discovered a distinct population of learning-related m6A- modified RNAs at the synapse, which includes the long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 (Malat1). RNA immunoprecipitation and mass spectrometry revealed 12 new synapse-specific learning-induced m6A readers in the mPFC of male C57/BL6 mice, with m6A-modified Malat1 binding to a subset of these, including CYFIP2 and DPYSL2. In addition, a cell type- and synapse-specific, and state-dependent, reduction of m6A on Malat1 impairs fear-extinction memory; an effect that likely occurs through a disruption in the interaction between Malat1 and DPYSL2 and an associated decrease in dendritic spine formation. These findings highlight the critical role of m6A in regulating the functional state of RNA during the consolidation of fear-extinction memory, and expand the repertoire of experience-dependent m6A readers in the synaptic compartment.SIGNIFICANCE STATEMENT We have discovered that learning-induced m6A-modified RNA (including the long noncoding RNA, Malat1) accumulates in the synaptic compartment. We have identified several new m6A readers that are associated with fear extinction learning and demonstrate a causal relationship between m6A-modified Malat1 and the formation of fear-extinction memory. These findings highlight the role of m6A in regulating the functional state of an RNA during memory formation and expand the repertoire of experience-dependent m6A readers in the synaptic compartment.
Assuntos
Medo , RNA Longo não Codificante , Animais , Masculino , Camundongos , Extinção Psicológica , Medo/fisiologia , Aprendizagem/fisiologia , RNA Longo não Codificante/metabolismo , Sinapses/metabolismoRESUMO
Post-translational modifications including protein ubiquitination regulate a plethora of cellular processes in distinct manners. RNA N6-methyladenosine is the most abundant post-transcriptional modification on mammalian mRNAs and plays important roles in various physiological and pathological conditions including hematologic malignancies. We previously determined that the RNA N6-methyladenosine eraser ALKBH5 is necessary for the maintenance of acute myeloid leukemia (AML) stem cell function, but the post-translational modifications involved in ALKBH5 regulation remain elusive. Here, we show that deubiquitinase ubiquitin-specific peptidase 9X (USP9X) stabilizes ALKBH5 and promotes AML cell survival. Through the use of mass spectrometry as an unbiased approach, we identify USP9X and confirm that it directly binds to ALKBH5. USP9X stabilizes ALKBH5 by removing the K48-linked polyubiquitin chain at K57. Using human myeloid leukemia cells and a murine AML model, we find that genetic knockdown or pharmaceutical inhibition of USP9X inhibits leukemia cell proliferation, induces apoptosis, and delays AML development. Ectopic expression of ALKBH5 partially mediates the function of USP9X in AML. Overall, this study uncovers deubiquitinase USP9X as a key for stabilizing ALKBH5 expression and reveals the important role of USP9X in AML, which provides a promising therapeutic strategy for AML treatment in the clinic.
Assuntos
Homólogo AlkB 5 da RNA Desmetilase , Leucemia Mieloide Aguda , Ubiquitina Tiolesterase , Animais , Humanos , Camundongos , Homólogo AlkB 5 da RNA Desmetilase/genética , Linhagem Celular Tumoral , Sobrevivência Celular , Leucemia Mieloide Aguda/genética , RNA , Ubiquitina Tiolesterase/genética , UbiquitinaçãoRESUMO
BACKGROUND: The RNA m6A modification has been implicated in multiple neurological diseases as well as macrophage activation. However, whether it regulates microglial activation during hypoxic-ischemic brain damage (HIBD) in neonates remains unknown. Here, we aim to examine whether the m6A modification is involved in modulating microglial activation during HIBD. We employed an oxygen and glucose deprivation microglial model for in vitro studies and a neonatal mouse model of HIBD. The brain tissue was subjected to RNA-seq to screen for significant changes in the mRNA m6A regulator. Thereafter, we performed validation and bioinformatics analysis of the major m6A regulators. RESULTS: RNA-seq analysis revealed that, among 141 m6A regulators, 31 exhibited significant differential expression (FC (abs) ≥ 2) in HIBD mice. We then subjected the major m6A regulators Mettl3, Mettl14, Fto, Alkbh5, Ythdf1, and Ythdf2 to further validation, and the results showed that all were significantly downregulated in vitro and in vivo. GO analysis reveals that regulators are mainly involved in the regulation of cellular and metabolic processes. The KEGG results indicate the involvement of the signal transduction pathway. CONCLUSIONS: Our findings demonstrate that m6A modification of mRNA plays a crucial role in the regulation of microglial activation in HIBD, with m6A-associated regulators acting as key modulators of microglial activation.
Assuntos
Ativação de Macrófagos , Microglia , Animais , Camundongos , Animais Recém-Nascidos , Encéfalo , RNA Mensageiro/genéticaRESUMO
Bisphenol A (BPA) is a commonly used plastic additive. Since BPA has been banned in maternal and infant food containers in many countries, BPA substitutes have been widely introduced to replace it. By systematically assessing the potential developmental toxicity of BPA substitutes, we observed that the 41-150 nM in vivo BPC exposure (around the reported concentration detected in infant urine: 6-186 nM) induced cardiac defects in zebrafish. Mechanistically, BPC disrupted m6A homeostasis by downregulation of the key m6A methyltransferase, Mettl3, thereby causing the m6A reader, Igf2bp2b, to fail in recognizing and stabilizing the inefficiently m6A-modified acox1 and tnnt2d mRNA. Then, downregulation of Acox1 (a regulator in cardiac fatty acid metabolism) and Tnnt2d (a component of cardiac troponin for muscle contraction) led to cardiac defects. Indeed, the dual cardiac functional axes regulated by the same m6A reader in response to BPC provided new insight into the regulatory mechanisms of epitranscriptomics and cardiac development. Collectively, our study not only presented evidence showing that the internal exposure levels of BPC in humans could lead to cardiac developmental defects but also demonstrated the underlying mechanism of BPC-mediated defects by disrupting the Mettl3-m6A-Igf2bp2b-Acox1/Tnnt2d pathways, which provided potential molecular markers associated with BPC exposure.
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In eukaryotes, RNA N6-methyladenosine (m6A) modification and microRNA (miRNA)-mediated RNA silencing represent two critical epigenetic regulatory mechanisms. The m6A methyltransferase complex (MTC) and the microprocessor complex both undergo liquid-liquid phase separation to form nuclear membraneless organelles. Although m6A methyltransferase has been shown to positively regulate miRNA biogenesis, a mechanism of reciprocal regulation between the MTC and the microprocessor complex has remained elusive. Here, we demonstrate that the MTC and the microprocessor complex associate with each other through the METHYLTRANSFERASE B (MTB)-SERRATE (SE) interacting module. Knockdown of MTB impaired miRNA biogenesis by diminishing microprocessor complex binding to primary miRNAs (pri-miRNAs) and their respective MIRNA loci. Additionally, loss of SE function led to disruptions in transcriptome-wide m6A modification. Further biochemical assays and fluorescence recovery after photobleaching (FRAP) assay indicated that SE enhances the liquid-liquid phase separation and solubility of the MTC. Moreover, the MTC exhibited enhanced retention on chromatin and diminished binding to its RNA substrates in the se mutant background. Collectively, our results reveal the substantial regulatory interplay between RNA m6A modification and miRNA biogenesis.
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Conflict between genes inherited from the mother (matrigenes) and the father (patrigenes) is predicted to arise during social interactions among offspring if these genes are not evenly distributed among offspring genotypes. This intragenomic conflict drives parent-specific transcription patterns in offspring resulting from parent-specific epigenetic modifications. Previous tests of the kinship theory of intragenomic conflict in honey bees (Apis mellifera) provided evidence in support of theoretical predictions for variation in worker reproduction, which is associated with extreme variation in morphology and behavior. However, more subtle behaviors - such as aggression - have not been extensively studied. Additionally, the canonical epigenetic mark (DNA methylation) associated with parent-specific transcription in plant and mammalian model species does not appear to play the same role as in honey bees, and thus the molecular mechanisms underlying intragenomic conflict in this species is an open area of investigation. Here, we examined the role of intragenomic conflict in shaping aggression in honey bee workers through a reciprocal cross design and Oxford Nanopore direct RNA sequencing. We attempted to probe the underlying regulatory basis of this conflict through analyses of parent-specific RNA m6A and alternative splicing patterns. We report evidence that intragenomic conflict occurs in the context of honey bee aggression, with increased paternal and maternal allele-biased transcription in aggressive compared to non-aggressive bees, and higher paternal allele-biased transcription overall. However, we found no evidence to suggest that RNA m6A or alternative splicing mediate intragenomic conflict in this species.
Assuntos
Agressão , RNA , Abelhas , Animais , Metilação de DNA , Alelos , Processamento Alternativo , MamíferosRESUMO
BACKGROUND: Although the relationship between type 2 diabetes (T2D) and the increased risk of colorectal carcinogenesis is widely defined in clinical studies, the therapeutic methods and molecular mechanism of T2D-induced colon cancer and how does hyperglycemia affect the progression is still unknown. Here, we studied the function of lactoferrin (LF) in suppressing the progression of colon cancer in T2D mice, and uncovered the related molecular mechanisms in DNA 5mC and RNA m6A levels. METHODS: We examined the effects of LF (50% iron saturation) on the migration and invasion of colon tumor cells under high concentration of glucose. Then, transcriptomics and DNA methylation profilings of colon tumor cells was co-analyzed to screen out the special gene (NT5DC3), and the expression level of NT5DC3 in 75 clinical blood samples was detected by q-PCR and western blot, to investigate whether NT5DC3 was a biomarker to distinguish T2D patients and T2D-induced colon cancer patients from healthy volunteers. Futhermore, in T2D mouse with xenografted colon tumor models, the inhibitory effects of LF and NT5DC3 protein on colon tumors were investigated. In addition, epigenetic alterations were measured to examine the 5mC/m6A modification sites of NT5DC3 regulated by LF. Utilizing siRNA fragments of eight m6A-related genes, the special gene (WTAP) regulating m6A of NT5DC was proved, and the effect of LF on WTAP/NT5DC3/HKDC1 axis was finally evaluated. RESULTS: A special gene NT5DC3 was screened out through co-analysis of transcriptomics and DNA methylation profiling, and HKDC1 might be a downstream sensor of NT5DC3. Mechanistically, LF-dependent cellular DNA 5mC and RNA m6A profiling remodeling transcriptionally regulate NT5DC3 expression. WTAP plays a key role in regulating NT5DC3 m6A modification and subsequently controls NT5DC3 downstream target HKDC1 expression. Moreover, co-treatment of lactoferrin and NT5DC3 protein restrains the growth of colon tumors by altering the aberrant epigenetic markers. Strikingly, clinical blood samples analysis demonstrates NT5DC3 protein expression is required to direct the distinction of T2D or T2D-induced colon cancer with healthy humans. CONCLUSIONS: Together, this study reveals that lactoferrin acts as a major factor to repress the progression of colon cancer under hyperglycemia, thus, significantly expanding the landscape of natural dietary mediated tumor suppression.
Assuntos
Neoplasias do Colo , Diabetes Mellitus Tipo 2 , Hiperglicemia , Humanos , Animais , Camundongos , Lactoferrina/genética , Neoplasias do Colo/genética , Hiperglicemia/complicações , Hiperglicemia/genética , Modelos Animais de Doenças , Fatores de Processamento de RNA , Proteínas de Ciclo Celular , HexoquinaseRESUMO
While arsenic or BaP alone exposure can cause lung cancer, studies showed that arsenic plus BaP co-exposure displays a significantly stronger lung tumorigenic effect. However, the underlying mechanism has not been well understood. Studies showed that RNA molecules are chemically modified. The most frequently occurring RNA modification in eukaryotic messenger RNAs is the N6-methyladenosine (m6A) methylation. This study aimed to determine whether arsenic plus BaP exposure alters RNA m6A methylation and its role in lung tumorigenic effect of arsenic plus BaP exposure. Human bronchial epithelial cells transformed by exposure to arsenic or BaP alone, and arsenic plus BaP and mouse xenograft tumorigenesis models were used in this study. It was found that arsenic plus BaP exposure-transformed cells have significantly higher levels of RNA m6A methylation than arsenic or BaP alone exposure-transformed human bronchial epithelial cells. Western blot analysis showed that arsenic plus BaP exposure greatly up-regulates the m6A writer methyltransferase like-3 (METTL3) expression levels in cultured cells and mouse lung tissues. METTL3 knockdown in cells transformed by arsenic plus BaP exposure drastically reduced their RNA m6A methylation levels. Functional studies revealed that METTL3 knockdown in cells transformed by arsenic plus BaP exposure greatly reduces their anchorage-dependent and -independent growth, cancer stem cell characters and tumorigenesis. The findings from this study suggest that arsenic plus BaP co-exposure causes epitranscriptomic dysregulation, which may contribute significantly to arsenic plus BaP co-exposure-caused synergistic lung tumorigenic effect.
Assuntos
Arsênio , Metiltransferases , Células-Tronco Neoplásicas , RNA , Animais , Humanos , Camundongos , Arsênio/toxicidade , Arsênio/metabolismo , Benzo(a)pireno/metabolismo , Benzo(a)pireno/toxicidade , Carcinogênese/induzido quimicamente , Carcinogênese/genética , Metiltransferases/genética , Metiltransferases/metabolismo , Células-Tronco Neoplásicas/metabolismo , Regulação para CimaRESUMO
Cancer cells respond to various stressful conditions through the dynamic regulation of RNA m6A modification. Doxorubicin is a widely used chemotherapeutic drug that induces DNA damage. It is interesting to know whether cancer cells regulate the DNA damage response and doxorubicin sensitivity through RNA m6A modification. Here, we found that doxorubicin treatment significantly induced RNA m6A methylation in breast cancer cells in both a dose- and a time-dependent manner. However, protein arginine methyltransferase 5 (PRMT5) inhibited RNA m6A modification under doxorubicin treatment by enhancing the nuclear translocation of the RNA demethylase AlkB homolog 5 (ALKBH5), which was previously believed to be exclusively localized in the nucleus. Then, ALKBH5 removed the m6A methylation of BRCA1 for mRNA stabilization and further enhanced DNA repair competency to decrease doxorubicin efficacy in breast cancer cells. Importantly, we identified the approved drug tadalafil as a novel PRMT5 inhibitor that could decrease RNA m6A methylation and increase doxorubicin sensitivity in breast cancer. The strategy of targeting PRMT5 with tadalafil is a promising approach to promote breast cancer sensitivity to doxorubicin through RNA methylation regulation.
Assuntos
Neoplasias da Mama , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Desmetilação , Doxorrubicina/farmacologia , Feminino , Humanos , Proteína-Arginina N-Metiltransferases/genética , RNA , TadalafilaRESUMO
Circular RNAs (circRNAs) are a class of endogenous functional RNA generated by back-splicing. Recently, circRNAs have been found to have certain coding potential. Proteins/peptides translated from circRNAs play essential roles in various diseases. Here, we briefly summarize the basic knowledge and technologies that are usually applied to study circRNA translation. Then, we focus on the research progress of circRNA translation in cardiovascular diseases and discuss the perspective and future direction of translatable circRNA study in cardiovascular diseases.
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Honeybees play a crucial role as pollinators for crops and are regarded as sensitive bioindicators of environmental health. The widespread use of pesticides poses a severe threat to honeybee survival. However, there is limited information available on the specific risks associated with fipronil exposure in honeybees, particularly concerning the impact on RNA methylation throughout their lifespan. This study aimed to evaluate the effects of sublethal concentrations of fipronil on RNA m6A and m5C methylations, along with the associated genes in honeybee larvae and newly emerged adults. LC-MS/MS analysis revealed a notable hypomethylation of m5C in larvae, while hypermethylation of m6A was observed in the adult brain. Significant changes in the expression of genes such as AmWTAP, AmYTHDF, AmALKBH4, AmALKBH6, AmALKBH8, AmNSUN5, AmNOP2, AmTET1, and AmYBX1 were observed in the adult brain, whereas alterations in the expression of AmNSUN2, AmMETTL14, AmALKBH1, AmALKBH4, AmALKBH6 AmALYREF, AmTET1, and AmYBX1 were observed in the larvae. Notably, the expression of AmALKBH1 was not detected in any fipronil-treated larvae, suggesting its potential as an early risk indicator for honeybee larvae in future assessments. This pioneering study provides insights into the effects of fipronil on RNA methylations in honeybees and explores the possibility of employing RNA methylation as a tool for assessing pesticide risks in this important pollinator species. These findings offer new perspectives on honeybee protection and the development of toxicity evaluation systems for pesticides.
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The development of mammalian skeletal muscle is a highly complex process involving multiple molecular interactions. As a prevalent RNA modification, N6-methyladenosine (m6A) regulates the expression of target genes to affect mammalian development. Nevertheless, it remains unclear how m6A participates in the development of goat muscle. In this study, methyltransferase 3 (METTL3) was significantly enriched in goat longissimus dorsi (LD) tissue. In addition, the global m6A modification level and differentiation of skeletal muscle satellite cells (MuSCs) were regulated by METTL3. By performing mRNA-seq analysis, 8050 candidate genes exhibited significant changes in expression level after the knockdown of METTL3 in MuSCs. Additionally, methylated RNA immunoprecipitation sequencing (MeRIP-seq) illustrated that myocyte enhancer factor 2c (MEF2C) mRNA contained m6A modification. Further experiments demonstrated that METTL3 enhanced the differentiation of MuSCs by upregulating m6A levels and expression of MEF2C. Moreover, the m6A reader YTH N6-methyladenosine RNA binding protein C1 (YTHDC1) was bound and stabilized to MEF2C mRNA. The present study reveals that METTL3 enhances myogenic differentiation in MuSCs by regulating MEF2C and provides evidence of a post-transcriptional mechanism in the development of goat skeletal muscle.
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Heterosis has been extensively utilized to increase productivity in crops, yet the underlying molecular mechanisms remain largely elusive. Here, we generated transcriptome-wide profiles of mRNA abundance, m6A methylation, and translational efficiency from the maize F1 hybrid B73×Mo17 and its two parental lines to ascertain the contribution of each regulatory layer to heterosis at the seedling stage. We documented that although the global abundance and distribution of m6A remained unchanged, a greater number of genes had gained an m6A modification in the hybrid. Superior variations were observed at the m6A modification and translational efficiency levels when compared with mRNA abundance between the hybrid and parents. In the hybrid, the vast majority of genes with m6A modification exhibited a non-additive expression pattern, the percentage of which was much higher than that at levels of mRNA abundance and translational efficiency. Non-additive genes involved in different biological processes were hierarchically coordinated by discrete combinations of three regulatory layers. These findings suggest that transcriptional and post-transcriptional regulation of gene expression make distinct contributions to heterosis in hybrid maize. Overall, this integrated multi-omics analysis provides a valuable portfolio for interpreting transcriptional and post-transcriptional regulation of gene expression in hybrid maize, and paves the way for exploring molecular mechanisms underlying hybrid vigor.