BACKGROUND: Fibrosis of the connective tissue in the vaginal wall predominates in pelvic organ prolapse (POP), which is characterized by excessive fibroblast-to-myofibroblast differentiation and abnormal deposition of the extracellular matrix (ECM). Our study aimed to investigate the effect of ECM stiffness on vaginal fibroblasts and to explore the role of methyltransferase 3 (METTL3) in the development of POP. METHODS: Polyacrylamide hydrogels were applied to create an ECM microenvironment with variable stiffness to evaluate the effects of ECM stiffness on the proliferation, differentiation, and expression of ECM components in vaginal fibroblasts. METTL3 small interfering RNA and an overexpression vector were transfected into vaginal fibroblasts to evaluate the effects of METTL3 silencing and overexpression on matrix stiffness-induced vaginal fibroblast-to-myofibroblast differentiation and abnormal modulation of the ECM. Both procedures were detected by 5-ethynyl-2'-deoxyuridine (EdU) staining, Western blotting (WB), quantitative real-time polymerase chain reaction (RT-qPCR), and immunofluorescence (IF). RESULTS: Vaginal fibroblasts from POP patients exhibited increased proliferation ability, increased expression of α-smooth muscle actin (α-SMA), decreased expression of collagen I/III, and significantly decreased expression of tissue inhibitors of matrix metalloproteinases (TIMPs) in the stiff matrix ( P <0.05). Compared with those from non-POP patients, vaginal wall tissues from POP patients demonstrated a significant increase in METTL3 content ( P <0.05). However, silencing METTL3 expression in vaginal fibroblasts with high ECM stiffness resulted in decreased proliferation ability, decreased α-SMA expression, an increased ratio of collagen I/III, and increased TIMP1 and TIMP2 expression. Conversely, METTL3 overexpression significantly promoted the process of increased proliferation ability, increased α-SMA expression, decreased ratio of collagen I/III and decreased TIMP1 and TIMP2 expression in the soft matrix ( P <0.05). CONCLUSIONS Elevated ECM stiffness can promote excessive proliferation, differentiation, and abnormal ECM modulation, and the expression of METTL3 plays an important role in alleviating or aggravating matrix stiffness-induced vaginal fibroblast-to-myofibroblast differentiation and abnormal ECM modulation.
Humans ; Female ; Extracellular Matrix/metabolism* ; Cell Differentiation/genetics* ; Methyltransferases/metabolism* ; Pelvic Organ Prolapse/pathology* ; Fibroblasts/metabolism* ; Myofibroblasts/metabolism* ; Vagina/metabolism* ; Cell Proliferation/physiology* ; Cells, Cultured ; Middle Aged

OBJECTIVE: To detect the expression of methyltransferase-like 7B ( METTL7B) in bone marrow specimens of patients with acute myeloid leukemia (AML), and to analyze its influence and significance on clinical diagnosis, treatment, and prognosis of AML patients. METHODS: Bone marrow specimens from 60 newly diagnosed AML patients were collected as the observation group, and bone marrow specimens from 20 iron-deficiency anemia (IDA) patients were collected as the control group. Clinical and pathological data of AML patients were also collected. Real-time fluorescent quantitative PCR (qRT-PCR) was used to detect the expression of METTL7B in AML patients and IDA patients. Statistical analyses were performed to investigate the relationship between the expression level of METTL7B and clinical-pathological characteristics in AML patients, as well as the impact of METTL7B expression level on efficacy. Kaplan-Meier method was used to analyze the effect of METTL7B expression level on the overall survival time (OS) in AML patients. Meanwhile, a Cox proportional hazards regression model was constructed to identify the factors potentially affecting the prognosis of AML patients. RESULTS: Compared with the control group, the expression level of METTL7B was significantly upregulated in AML patients (P < 0.05). Compared with the low-expression group of METTL7B, the high-expression group had a higher proportion of patients with high white blood cell (WBC) count, poor prognosis, and ineffective treatment, and the differences were statistically significant (P < 0.05). The OS of patients in the high-expression group of METTL7B was significantly shorter than that in the low-expression group (P < 0.05). Multivariate Cox regression analysis showed that high WBC count, poor prognosis in prognosis stratification, and high expression of METTL7B were independent risk factors for the prognosis of AML patients (P < 0.05). CONCLUSION METTL7B is highly expressed in AML patients, and patients with high METTL7B expression exhibit shorter survival and poor prognosis. METTL7B is expected to serve as a new indicator for evaluating the prognosis of AML patients and may develop into a potential target for targeted treatment of AML in the future.
Humans ; Leukemia, Myeloid, Acute/metabolism* ; Prognosis ; Methyltransferases/metabolism* ; Male ; Female ; Middle Aged ; Adult ; Proportional Hazards Models

OBJECTIVES: To evaluate the effect of eye acupuncture on neural function and angiogenesis of ischemic cerebral tissue in rats, and explore the roles of METTL3-mediated m⁶A methylation and the HIF-1α/VEGF-A signal axis in mediating this effect. METHODS: Fifty SD rats were randomized into normal control group, sham-operated group, model group, eye acupuncture group and DMOG (a HIF-1α agonist) group. Rat models of cerebral ischemia/reperfusion injury (CIRI) were established using a modified thread thrombus method, and the changes in neurological deficits of the rats after interventions were evaluated. TTC and Nissl staining were used to examine the changes in infarction size and neuronal injury, and cerebral angiogenesis was detected by double-immunofluorescence staining. m⁶A methylation modification level in the brain tissue was detected by ELISA, and RT-qPCR and Western blotting were used to detect the mRNA and protein expressions of METTL3 and HIF-1α/VEGF-A. RESULTS: Compared with the control and sham-operated rats, the CIRI rats had significantly higher neurological deficit scores with larger cerebral infarction area, a greater number of CD31- and EDU-positive new vessels, higher expression levels of HIF-1α and VEGF-A, reduced number of Nissl bodies and m6A methylation level, and lowered METTL3 protein and mRNA expressions. All these changes were significantly improved by interventions with eye acupuncture after modeling or intraperitoneal injections of DMOG for 7 consecutive days prior to modeling, and the effects of the two interventions were similar. CONCLUSIONS Eye acupuncture can improve neurological deficits in CIRI rat models possibly by promoting cortical angiogenesis via upregulating METTL3-mediated m⁶A methylation and regulating the HIF-1α/VEGF-A signal axis.
Animals ; Rats, Sprague-Dawley ; Methyltransferases/metabolism* ; Reperfusion Injury/physiopathology* ; Methylation ; Hypoxia-Inducible Factor 1, alpha Subunit/metabolism* ; Rats ; Vascular Endothelial Growth Factor A/metabolism* ; Brain Ischemia/metabolism* ; Acupuncture Therapy ; Male ; Up-Regulation ; Neovascularization, Physiologic ; Angiogenesis ; Adenosine/analogs & derivatives*

Hypoxemia is a common pathological state characterized by low oxygen saturation in the blood. This condition compromises mucosal barrier integrity particularly in the gut and oral cavity. However, the mechanisms underlying this association remain unclear. This study used periodontitis as a model to investigate the role of platelet activation in oral mucosal immunopathology under hypoxic conditions. Hypoxia upregulated methyltransferase-like protein 4 (METTL4) expression in platelets, resulting in N⁶-methyladenine modification of mitochondrial DNA (mtDNA). This modification impaired mitochondrial transcriptional factor A-dependent cytosolic mtDNA degradation, leading to cytosolic mtDNA accumulation. Excess cytosolic mt-DNA aberrantly activated the cGAS-STING pathway in platelets. This resulted in excessive platelet activation and neutrophil extracellular trap formation that ultimately exacerbated periodontitis. Targeting platelet METTL4 and its downstream pathways offers a potential strategy for managing oral mucosa immunopathology. Further research is needed to examine its broader implications for mucosal inflammation under hypoxic conditions.
DNA, Mitochondrial/metabolism* ; Mouth Mucosa/pathology* ; Hypoxia/immunology* ; Methyltransferases/metabolism* ; Blood Platelets/metabolism* ; Animals ; Periodontitis/immunology* ; Humans ; Platelet Activation ; Mice

Diabetic cardiomyopathy (DCM) is a medical condition characterized by cardiac remodeling and dysfunction in individuals with diabetes mellitus. Sarcoplasmic reticulum (SR) and mitochondrial Ca²⁺ overload in cardiomyocytes have been recognized as biological hallmarks in DCM; however, the specific factors underlying these abnormalities remain largely unknown. In this study, we aimed to investigate the role of a cardiac-specific long noncoding RNA, D830005E20Rik (Trdn-as), in DCM. Our results revealed the remarkably upregulation of Trdn-as in the hearts of the DCM mice and cardiomyocytes treated with high glucose (HG). Knocking down Trdn-as in cardiac tissues significantly improved cardiac dysfunction and remodeling in the DCM mice. Conversely, Trdn-as overexpression resulted in cardiac damage resembling that observed in the DCM mice. At the cellular level, Trdn-as induced Ca²⁺ overload in the SR and mitochondria, leading to mitochondrial dysfunction. RNA-seq and bioinformatics analyses identified calsequestrin 2 (Casq2), a primary calcium-binding protein in the junctional SR, as a potential target of Trdn-as. Further investigations revealed that Trdn-as facilitated the recruitment of METTL14 to the Casq2 mRNA, thereby enhancing the m⁶A modification of Casq2. This modification increased the stability of Casq2 mRNA and subsequently led to increased protein expression. When Casq2 was knocked down, the promoting effects of Trdn-as on Ca²⁺ overload and mitochondrial damage were mitigated. These findings provide valuable insights into the pathogenesis of DCM and suggest Trdn-as as a potential therapeutic target for this condition.
Animals ; Diabetic Cardiomyopathies/pathology* ; RNA, Long Noncoding/genetics* ; Myocytes, Cardiac/metabolism* ; Mice ; Calsequestrin/genetics* ; Calcium/metabolism* ; Male ; Sarcoplasmic Reticulum/metabolism* ; Methyltransferases/metabolism* ; Mice, Inbred C57BL ; Mitochondria, Heart/metabolism* ; Disease Models, Animal ; Mitochondria/metabolism*

N⁶-methyladenosine (m⁶A) modification plays a critical role in cell cycle regulation, while the mechanism of m⁶A in regulating mitosis remains underexplored. Here, we found that the total m⁶A modification level in cells increased during mitosis by the liquid chromatography-mass spectrometry/mass spectrometry and m⁶A dot blot assays. Silencing methyltransferase-like 3 (METTL3) or METTL14 results in delayed mitosis, abnormal spindle assembly, and chromosome segregation defects by the immunofluorescence. By analyzing transcriptome-wide m⁶A targets in HeLa cells, we identified polo-like kinase 1 (PLK1) as a key gene modified by m⁶A in regulating mitosis. Specifically, through immunoblotting and RNA pulldown, m⁶A modification inhibits PLK1 translation via YTH N⁶-methyladenosine RNA binding protein 1, thus mediating cell cycle homeostasis. Demethylation of PLK1 mRNA leads to significant mitotic abnormalities. These findings highlight the critical role of m⁶A in regulating mitosis and the potential of m⁶A as a therapeutic target in proliferative diseases such as cancer.
Humans ; Polo-Like Kinase 1 ; Cell Cycle Proteins/metabolism* ; Proto-Oncogene Proteins/metabolism* ; Protein Serine-Threonine Kinases/metabolism* ; Mitosis/physiology* ; HeLa Cells ; Adenosine/genetics* ; Methyltransferases/metabolism* ; RNA, Messenger/metabolism* ; RNA-Binding Proteins/metabolism*

Geminiviruses cause substantial crop yield losses worldwide. The replication initiator protein (Rep) encoded by geminiviruses is indispensable for geminiviral replication. The Rep protein encoded by beet severe curly top virus (BSCTV, genus Curtovirus, family Geminiviridae) induces VARIANT IN METHYLATION 5 (VIM5) expression in Arabidopsis leaves upon BSCTV infection. VIM5 functions as a ubiquitination-related E3 ligase to promote the proteasomal degradation of methyltransferases, resulting in reduction of methylation levels in the BSCTV C2-3 promoter. However, the specific domains of Rep responsible for VIM5 induction remain poorly characterized. Although Rep proteins from several geminiviruses act as viral suppressors of RNA silencing (VSRs), whether BSCTV Rep also possesses VSR activity remains to be illustrated. In this study, we employed a transient expression system in the 16c-GFP transgenic and the wild-type Nicotiana benthamiana plants to analyze the VSR and the VIM5-inducing activities of different truncated Rep proteins haboring distinct domains. We found that the N-terminal domain (amino acids 1-180) of Rep suppressed GFP silencing in 16c-GFP transgenic N. benthamiana leaves. The minimal N-terminal fragment (amino acids 1-104) induced VIM5 expression upon co-infiltration, while C-terminal truncations lacked VIM5-inducing activity. Our results indicate that the N-terminal domain of Rep encoded by BSCTV mediates the suppression of RNA silencing and induces VIM5 expression. Thus, our findings contribute to a better understanding of interactions between geminiviral Rep and plant hosts.
Geminiviridae/genetics* ; Nicotiana/metabolism* ; Arabidopsis/metabolism* ; RNA Interference ; Viral Proteins/metabolism* ; Arabidopsis Proteins/metabolism* ; Plants, Genetically Modified/metabolism* ; Protein Domains ; Plant Diseases/virology* ; Methyltransferases/metabolism* ; Ubiquitin-Protein Ligases/metabolism* ; DNA Helicases/genetics*

BACKGROUND: Lung cancer is a major threat to human health. The molecular mechanisms related to the occurrence and development of lung cancer are complex and poorly known. Exploring molecular markers related to the development of lung cancer is helpful to improve the effect of early diagnosis and treatment. Long non-coding RNA (lncRNA) THAP7-AS1 is known to be highly expressed in gastric cancer, but has been less studied in other cancers. The aim of the study is to explore the role and mechanism of methyltransferase-like 3 (METTL3) mediated up-regulation of N6-methyladenosine (m6A) modified lncRNA THAP7-AS1 expression in promoting the development of lung cancer. METHODS: Samples of 120 lung cancer and corresponding paracancerous tissues were collected. LncRNA microarrays were used to analyze differentially expressed lncRNAs. THAP7-AS1 levels were detected in lung cancer, adjacent normal tissues and lung cancer cell lines by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The diagnostic value of THAP7-AS1 in lung cancer and the relationship between THAP7-AS1 expression and survival rate and clinicopathological parameters were analyzed. Bioinformatics analysis, methylated RNA immunoprecipitation (meRIP), RNA pull-down and RNA-immunoprecipitation (RIP) assay were used to investigate the molecular regulation mechanism of THAP7-AS1. Cell proliferation, migration, invasion and tumorigenesis of SPC-A-1 and NCI-H1299 cells were determined by MTS, colony-formation, scratch, Transwell and xenotransplantation in vivo, respectively. Expression levels of phosphoinositide 3-kinase/protein kenase B (PI3K/AKT) signal pathway related protein were detected by Western blot. RESULTS: Expression levels of THAP7-AS1 were higher in lung cancer tissues and cell lines (P<0.05). THAP7-AS1 has certain diagnostic value in lung cancer [area under the curve (AUC)=0.737], and its expression associated with overall survival rate, tumor size, tumor-node-metastasis (TNM) stage and lymph node metastasis (P<0.05). METTL3-mediated m6A modification enhanced THAP7-AS1 expression. The cell proliferation, migration, invasion and the volume and mass of transplanted tumor were all higher in the THAP7-AS1 group compared with the NC group and sh-NC group of SPC-A-1 and NCI-H1299 cells, while the cell proliferation, migration and invasion were lower in the sh-THAP7-AS1 group (P<0.05). THAP7-AS1 binds specifically to Cullin 4B (CUL4B). The cell proliferation, migration, invasion, and expression levels of phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), phosphoinositide-3 kinase, catalytic subunit delta (PIK3CD), phospho-phosphatidylinositol 3-kinase (p-PI3K), phospho-protein kinase B (p-AKT) and phospho-mammalian target of rapamycin (p-mTOR) were higher in the THAP7-AS1 group compared with the Vector group of SPC-A-1 and NCI-H1299 cells (P<0.05). CONCLUSIONS LncRNA THAP7-AS1 is stably expressed through m6A modification mediated by METTL3, and combines with CUL4B to activate PI3K/AKT signal pathway, which promotes the occurrence and development of lung cancer.
Humans ; Lung Neoplasms/pathology* ; RNA, Long Noncoding/metabolism* ; Phosphatidylinositol 3-Kinases/metabolism* ; Up-Regulation ; Proto-Oncogene Proteins c-akt/metabolism* ; Cell Line, Tumor ; Cell Proliferation/genetics* ; Gene Expression Regulation, Neoplastic ; Methyltransferases/metabolism* ; Cullin Proteins/genetics*

Humans ; Cardiovascular Diseases ; Enhancer of Zeste Homolog 2 Protein/metabolism* ; Histones ; Methyltransferases

Humans ; Cardiovascular Diseases ; Enhancer of Zeste Homolog 2 Protein/metabolism* ; Histones ; Methyltransferases