Upregulation of circGDI2 inhibits tumorigenesis by stabilizing the expression of RNA m6A demethylase FTO in oral squamous cell carcinoma.

Background: Oral squamous cell carcinoma (OSCC) is a malignant tumour that is difficult to identify and prone to metastasis and invasion. Circular RNAs (circRNAs) are important cancer regulators and can be used as potential biomarkers. However, OSCC-related circRNAs need to be further explored. We investigated the role of circGDI2 in OSCC and explored its downstream regulatory mechanisms. Methods: Quantitative real-time PCR was used to detect the expression levels of circGDI2 and fat mass and obesity-associated protein (FTO) in cells. Lentiviral transfection was used to construct stable circGDI2 overexpressing cells for subsequent cell function tests. RNA pull-down, RNA Immunoprecipitation (RIP), western blotting, and protein stability assays were conducted to detect circGDI2 binding proteins and their functions. CCK8, Transwell, and wound healing assays were used to verify cell functions after overexpressing circGDI2 or suppressing FTO expression. Animal experiments were performed to verify the results in vivo. Results: The expression of circGDI2 was markedly decreased in both OSCC cell lines and patient tissues. Overexpression of circGDI2 in OSCC cell lines led to decreased proliferation, migration, and invasion abilities. Knockdown of circGDI2 showed the opposite trend. CircGDI2 has been validated to interact with the FTO protein within cells, as evidenced by mass spectrometry and RIP assays. This interaction was found to prevent the degradation of the FTO protein. Dot blot analysis showed a reduction in N6-methyladenosine (m6A) modification after circGDI2 overexpression. Reduced FTO levels reversed the inhibitory effects of circGDI2 overexpression on cell proliferation, migration, and invasion in vitro and on tumorigenesis in vivo. Conclusions: CircGDI2 functions as a tumour suppressor by binding to the FTO protein to reduce RNA m6A modification levels and ultimately inhibit proliferation and migration in OSCC cells. This study indicates the potential use of circGDI2 as a new target for the prevention and treatment of OSCC.

Fat mass and obesity-related protein contributes to the development and maintenance of bone cancer pain in rats by abrogating m6A methylation of RNA.

Effective prevention and treatment options for bone cancer-related pain (BCP) are lacking. In recent years, numerous studies have investigated the association between m6A epigenetic modifications and pain, revealing their significant role in pain initiation and maintenance. This study aimed to provide theoretical support for the treatment of BCP and to identify target drugs for future development. Specifically, we investigated the involvement of fat mass and obesity-related protein (FTO) in rat models of BCP by administering varying doses (1/5/10 mg/kg) of the FTO inhibitor meclofenamic acid (MA) and assessing changes in mechanical sensitivity through domain analysis, gait analysis, and open-field experiments. After successfully establishing the BCP model, we verified it by performing mechanical sensitivity assessments. We observed significantly increased expression levels of the demethylase FTO within the spinal dorsal horn accompanied by decreased m6A methylation levels in the model. Compared with untreated BCP rats, remarkably improved behavioral responses indicative of reduced pain were observed in the model rats after administration of 10 mg/kg MA, concomitant with decreased expression levels of FTO and increased m6A methylation levels. Compared with untreated BCP rats, the expression levels of p-ERK and pro-inflammatory cytokines were also significantly decreased after MA administration. Taken together, FTO can downregulate m6A methylation level and activate ERK/inflammatory cytokines signaling pathway to maintain BCP in rats.

The melatonin-FTO-ATF4 signaling pathway protects granulosa cells from cisplatin-induced chemotherapeutic toxicity by suppressing ferroptosis.

PURPOSE: In cisplatin-induced premature ovarian failure (POF) mice, granulosa cells showed a high level of ferroptosis. Previous research has indicated that the fat mass and obesity-associated protein/activating transcription factor 4 (FTO/ATF4) axis was involved in the regulation of ferroptosis. The purpose of this study was to explore the role of the FTO/ATF4 axis in cisplatin-induced ferroptosis in granulosa cell. METHODS: The extent of ferroptosis was assessed by transmission electron microscopy (TEM) and ROS, GPX, GSH, and MDA assays. Western blotting was used to evaluate the protein expression levels of ferroptosis-related molecules. Ferroptosis activator and inhibitor were also used. RESULTS: We found that ferroptosis increased in a concentration-dependent manner in cisplatin-induced injured granulosa cells, accompanied by the downregulation of FTO. In addition, gain- and loss-of-function studies showed that FTO affects ferroptosis in injured cells by regulating ATF4 expression. Ferrostatin-1 inhibited the effect of FTO downregulation on injured granulosa cells ferroptosis, and erastin reversed the protective effect of FTO on ferroptosis in injured granulosa cells. Finally, melatonin was used, and we found that melatonin reduced ferroptosis in cisplatin-induced injured granulosa cells by upregulating FTO expression. CONCLUSION: Our study demonstrated that cisplatin induced granulosa cell ferroptosis by downregulating the expression of FTO. ATF4 was identified as a downstream target of FTO, and overexpression of ATF4 reversed the effects of decreased FTO on ferroptosis. Additionally, melatonin mitigates the cytotoxic effects of cisplatin by upregulating FTO expression. The melatonin-FTO-ATF4 signaling pathway plays a vital role in the treatment of cisplatin-induced POF.

FTO/m6A mediates miR-138-5p maturation and regulates gefitinib resistance of lung adenocarcinoma cells by miR-138-5p/LCN2 axis.

BACKGROUND: Lung cancer (LC) occupies an important position in the lethality of cancer patients. Acquired resistance to gefitinib in lung adenocarcinoma (LUAD) seriously affects the therapeutic efficacy of LC. Thus, it is of major scientific and clinical significance to probe the mechanism of gefitinib resistance in LUAD for ameliorating the prognosis of patients. METHODS: The expression of miRNAs in gefitinib-resistant LUAD cells was validated using qRT-PCR. Cell viability was assessed through CCK-8, whereas cell death was examined through PI staining. Changes in the ferroptosis process were evaluated by detecting the intracellular Glutathione (GSH), Malondialdehyde (MDA), and Reactive Oxygen Species (ROS) levels. Downstream targets of miR-138-5p were verified via luciferase reporter and RNA pull-down assays. RIP and qRT-PCR were employed to evaluate pri-miR-138-5p binding to DiGeorge critical region 8 (DGCR8) and the pri-miR-138-5p m6A modification level. Additionally, the impact of fat mass and obesity-associated protein (FTO) on LUAD gefitinib sensitivity was assessed in vivo by constructing a xenograft model. RESULTS: We observed that miR-138-5p was notably diminished in gefitinib-resistant cells. Overexpression of miR-138-5p suppressed viability while facilitated cell death and intracellular ferroptosis in gefitinib-resistant cells. Moreover, lipocalin 2 (LCN2) was the downstream target of miR-138-5p. The biological functions of miR-138-5p on gefitinib-resistant cells was reversed by introduction of LCN2. FTO suppressed the binding of DGCR8 to pri-miR-138-5p through m6A modification, thereby restraining the processing of miR-138-5p. Meanwhile, silencing of FTO enhanced the sensitivity of LUAD to gefitinib treatment. CONCLUSION: FTO suppressed the processing of miR-138-5p and then modulated the proliferation, death, and ferroptosis of gefitinib-resistant cells through the miR-138-5p/LCN2 pathway, which may put forward novel insights for clinically ameliorating the therapeutic effect of gefitinib in LUAD.

FTO mediates the diabetic kidney disease progression through regulating the m6A modification of NLRP3.

BACKGROUND: The objective of our research was to investigate the specific mechanism of FTO in diabetic kidney disease (DKD) progression. METHODS: The DKD model was established with renal tubular epithelial HK-2 cells and mice in vitro and in vivo. The N6-methyladenosine (m6A) content in cells was detected using dot plot assay and the m6A levels of NLRP3 was detected with the MeRIP assay. The mRNA and protein levels were tested with real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) and western blot. The IL-1ß and IL-18 levels were assessed with enzyme-linked immunosorbent assay (ELISA). The cell viability was measured by cell counting kit (CCK)-8 assay and cell pyroptosis was determined with Annexin V and propidium iodide (PI) double staining followed by flow cytometry analysis. RNA-binding protein immunoprecipitation (RIP) and dual luciferase reporter assays were conducted to detect the interaction between FTO and NLRP3. m6A levels were detected by Me-RIP assay. The renal injury was measured by observing the renal morphology and urine and blood levels of relevant indicators. RESULTS: The results indicated that high glucose treatment induced HK-2 cell pyroptosis. m6A levels were prominently elevated in high glucose treated HK-2 cells while FTO expression were significantly down-regulated. FTO over-expression promoted cell viability but inhibited pyroptosis of HK-2 cells under high glucose (HG) treatment. Moreover, FTO could inhibit NLRP3 expression. RIP and Me-RIP assays indicated that FTO could bind with NLRP3 and regulate its m6A modification level. Further luciferase assay confirmed that FTO binds with the 233-237 bp region of NLRP3. NLRP3 neutralized the function of FTO in the HG stimulated HK-2 cells. In vivo, the H&E staining showed that FTO over-expression alleviated the kidney injury and suppressed the pyroptosis induced by DKD. CONCLUSION: We found that FTO could inhibit the DKD progression in vivo and in vitro by regulated the m6A modification of NLRP3.

Genetic and Anthropometric Interplay: How Waist-to-Hip Ratio Modulates LDL-c Levels in Mexican Population.

BACKGROUND/OBJECTIVES: Genetic factors contribute to the physiopathology of obesity and its comorbidities. This study aimed to investigate the association of the SNPs ABCA1 (rs9282541), ADIPOQ (rs2241766), FTO (rs9939609), GRB14 (rs10195252), and LEPR (rs1805134) with various clinical, anthropometric, and biochemical variables. METHODS: The study included 396 Mexican mestizo individuals with obesity and 142 individuals with normal weight. Biochemical markers were evaluated from peripheral blood samples, and SNP genotyping was performed using PCR with TaqMan probes. A genetic risk score (GRS) was computed using an additive model. RESULTS: No significant associations were found between the SNPs ABCA1, ADIPOQ, FTO, and LEPR with obesity. However, the T allele of the GRB14 SNP was significantly associated with obesity (χ2 = 5.93, p = 0.01; OR = 1.52; 95% CI: 1.08-2.12). A multivariate linear regression model (adjusted R-squared: 0.1253; p < 0.001) predicting LDL-c levels among all participants (n = 538) identified significant (p < 0.05) beta coefficients for several anthropometric and biochemical variables, as well as for the GRS. Additionally, the interaction between the GRS and the waist-to-hip ratio (WHR) showed a negative beta coefficient (BC = -26.5307; p = 0.014). Participants with a WHR < 0.839 showed no effect of GRS on LDL-c concentration, while those with a WHR > 0.839 exhibited a greater effect of GRS (~9) at lower LDL-c concentrations (~50 mg/dL) and a lesser effect of GRS (~7) at higher LDL-c concentrations (~250 mg/dL). CONCLUSIONS: A significant interaction between genetics and WHR influences LDL-c in Mexicans, which may contribute to the prevention and clinical management of dyslipidemia and cardiovascular disease.

The N6-methyladenosine landscape of ovarian development and aging highlights the regulation by RNA stability and chromatin state.

The versatile epigenetic modification known as N6-methyladenosine (m6A) has been demonstrated to be pivotal in numerous physiological and pathological contexts. Nonetheless, the precise regulatory mechanisms linking m6A to histone modifications and the involvement of transposable elements (TEs) in ovarian development and aging are still not completely understood. First, we discovered that m6A modifications are highly expressed during ovarian aging (OA), with significant contributions from decreased m6A demethylase FTO and overexpressed m6A methyltransferase METTL16. Then, using FTO knockout mouse model and KGN cell line, we also observed that FTO deletion and METTL16 overexpression significantly increased m6A levels. This led to the downregulation of the methyltransferase SUV39H1, resulting in reduced H3K9me3 expression. The downregulation of SUV39H1 and H3K9me3 primarily activated LTR7 and LTR12, subsequently activating ERV1. This resulted in a decrease in cell proliferation, while the levels of apoptosis, cellular aging markers, and autophagy markers significantly increased in OA. In summary, our study offers intriguing insights into the role of m6A in regulating DNA epigenetics, including H3K9me3 and TEs, as well as autophagy, thereby accelerating OA.

Diet-induced Obesity: Pathophysiology, Consequences and Target Specific Therapeutic Strategies.

Diet has emerged as a pivotal factor in the current time for diet-induced obesity (DIO). A diet overloaded with fats and carbohydrates and unhealthy dietary habits contribute to the development of DIO through several mechanisms. The prominent ones include the transition of normal gut microbiota to obese microbiota, under-expression of AMPK, and abnormally high levels of adipogenesis. DIO is the root of many diseases. The present review deals with various aspects of DIO and its target proteins that can be specifically used for its treatment. Also, the currently available treatment strategies have been explored. It was found that the expression of five proteins, namely, PPARγ, FTO, CDK4, 14-3-3 ζ protein, and Galectin-1, is upregulated in DIO. They can be used as potential targets for drug-designing studies. Thus, with these targets, the treatment strategy for DIO using natural bioactive compounds can be a safer alternative to medications and bariatric surgeries.

Fat Mass and Obesity-Related (FTO) Gene Variant Is a Predictor of CVD in T2DM Patients.

Background: The role of the common FTO gene variant rs9939609 in obesity has been well established, and the FTO gene has a strong association with T2DM. Objective: To investigate the association of FTO gene variant rs9939609 with obesity-related parameters in T2DM and CVD patients. Materials and Methods: In this cross-sectional study, 280 subjects of either sex aged 45.10 ± 9.6 years were randomly divided into four groups, that is, T2DM, T2DM with CVD, nondiabetic with CVD disease, and normal control. These samples were genotyped by ARMS-PCR. The FTO gene association with obesity-related parameters in T2DM and CVD patients was analyzed by SPSS 22. Results: The TT genotype was the most common genotype (46.80%) in our study groups. The minor allele frequency (MAF) was significantly higher in T2DM patients (0.39 vs. 0.28), T2DM patients with CVD (0.43 vs. 0.28), and nondiabetic patients with CVD (0.35 vs. 0.28) as compared to control with p < 0.005. Carriers of the AA genotype of the FTO gene rs9939609 were significantly associated with increased BMI, WC, HbA1C, SBP, DBP, and TGs and lowered HDL cholesterol as compared to the TA and TT genotypes in T2DM and CVD patients with p < 0.005. The FTO gene variant rs9939609 showed a significant association with T2DM and CVD. The AA genotype odds ratio (OR) in T2DM was 1.48 (1.06-2.32), p = 0.006, and in CVD, it was 1.56 (1.04-2.4), p = 0.003. Conclusion: The FTO gene variant rs9939609 has a strong association with T2DM and CVD. The AA genotype of FTO gene variants rs9939609 showed a strong association with most of the risk factors of CVD and T2DM.

Genetic association of FTO gene polymorphisms with obesity and its related phenotypes: A case-control study.

Introduction: FTO gene belongs to the non-heme Fe (II) and 2 oxoglutarate-dependent dioxygenase superfamily. Polymorphisms within the first intron of the FTO gene have been examined across various populations, yielding disparate findings.The present study aimed to determine the impact of two intronic polymorphisms FTO 30685T/G (rs17817449) and -23525T/A (rs9939609) on the risk of obesity in Punjab, India. Methods: Genotypic and biochemical analysis were done for 671 unrelated participants (obese=333 and non-obese=338) (age≥18 years). Genotyping of the polymorphisms was done by PCR-RFLP method. However, 50% of the samples were sequenced by Sanger sequencing. Results: Both the FTO variants 30685 (TT vs GG: odds ratio (OR), 2.30; 95% confidence interval (CI), 1.39-3.79) and -23525 (TT vs AA: odds ratio (OR), 2.78; 95% confidence interval (CI), 1.37-5.64) showed substantial risk towards obesity by conferring it 2 times and 3 times, respectively. The analysis by logistic regression showed a significant association for both the variants 30685T/G (rs17817449) and -23525T/A (rs9939609) (OR=2.29; 95%CI: 1.47-3.57) and (OR=5.25; 95% CI: 2.68-10.28) under the recessive genetic model, respectively. The haplotype combination TA (30685; -23525) develops a 4 times risk for obesity (P=0.0001). Among obese, the G allele of 30685T/G and A- allele of -23525T/A showed variance in Body mass index (BMI), waist circumference (WC), waist-to-height ratio(WHtR), systolic blood pressure (SBP), diastolic blood pressure (DBP) and triglyceride(TG). Conclusion: The present investigation indicated that both the FTO 30685T/G (rs17817449) and -23525T/A (rs9939609) polymorphisms have a key impact on an individual's vulnerability to obesity in this population.

FTO diversely influences sensitivity of neuroblastoma cells to various chemotherapeutic drugs.

Chemotherapy resistance is a significant factor in treatment failure in patients with neuroblastoma (NB), and it directly affects patient prognosis. Therefore, identifying novel therapeutic targets to enhance chemosensitivity is essential to improve the cure rate and prognosis of patients with NB. In this study, we investigated the role of FTO in chemosensitivity of NB cells to various chemotherapeutic drugs. Our results showed that high FTO expression was positively correlated with increased survival probability and favorable prognostic factors in patients with NB. FTO overexpression inhibited cell proliferation, whereas FTO knockdown promoted cell proliferation in NB cells. FTO expression alteration had contrasting effects on NB cells' sensitivity to etoposide but had no significant impact on sensitivity to cisplatin. Downregulation of FTO reduced the sensitivity of NB cells to paclitaxel, whereas upregulation of FTO enhanced its sensitivity. Additionally, the sensitivities between patients with lower and higher FTO expression to various chemotherapeutic drugs or small-molecule inhibitors were different. Thus, FTO affects the sensitivities of NB cells differently depending on the different chemotherapeutic drugs and small-molecule inhibitors. This finding may guide physicians and patients choose the appropriate chemotherapeutic drugs or small-molecule inhibitors for treatment.

Comprehensive analysis of the expression patterns and function of the FTO-LINE1 axis in yak tissues and muscle satellite cells.

Background: The long interspersed nuclear element 1 (LINE1) retrotransposon has been identified as a specific substrate for fat mass and obesity-related gene (FTO), which facilitates the removal of N6-methyladenosine modifications from its targeted RNAs. Methods: This study examined the dynamic interaction between FTO and LINE1 in yak tissues and muscle satellite cells, utilizing RT-qPCR, RNA immunoprecipitation (RIP), immunofluorescence staining, and techniques involving overexpression and interference of FTO and LINE1 to elucidate the relationship between FTO and LINE1 in yak tissues and muscle satellite cells. Results: Cloning and analysis of the FTO coding sequence in Jiulong yak revealed a conserved protein structure across various Bos breeds, with notable homology observed with domestic yak, domestic cattle, and Java bison. Comprehensive examination of FTO and LINE1 gene expression patterns in Jiulong yaks revealed consistent trends across tissues in both sexes. FTO mRNA levels were markedly elevated in the heart and kidney, while LINE1 RNA was predominantly expressed in the heart. Immunoprecipitation confirmed the direct interaction between the FTO protein and LINE1 RNA in yak tissues and muscle satellite cells. The FTO-LINE1 axis was confirmed by a significant decrease in LINE1 RNA enrichment following its expression interference in yak muscle satellite cells. Overexpression of FTO substantially reduced the expression of recombinant myogenic factor 5 (MYF5). However, FTO interference had no discernible effect on MYF5 and myoblast determination protein 1 (MYOD1) mRNA levels. Immunofluorescence analysis revealed no alterations in Ki-67 protein expression following FTO interference or overexpression. However, phalloidin staining demonstrated enhancement in the myotube fusion rate of yak muscle satellite cells upon LINE1 interference. Conclusion: This comprehensive mapping of the FTO and LINE1 mRNA expression patterns establishes a direct interaction between the FTO protein and LINE1 RNA in yak. The findings suggest that FTO overexpression promotes muscle satellite cells differentiation, whereas LINE1 negatively regulates myotube fusion. The study provides fundamental insights into the role of the FTO-LINE1 axis in determining the fate of muscle satellite cells in yak, laying a solid theoretical foundation for future investigations.

The molecular regulated mechanism of METTL3 and FTO in lipid metabolism of Hu sheep.

BACKGROUND: Balanced lipid metabolism can improve the growth performance and meat quality of livestock. The m6A methylation-related genes METTL3 and FTO play important roles in animal lipid metabolism; however, the mechanism through which they regulate lipid metabolism in sheep is unclear. RESULTS: We established lipid deposition models of hepatocytes and preadipocytes in Hu sheep. In the hepatocyte lipid deposition model, the genes expression levels of FABP4, Accα, ATGL and METTL3, METTL14, and FTO-were significantly up-regulated after lipid deposition (P < 0.05). Transcriptomic and metabolomic analyses showed that lipid deposition had a significant effect on MAPK, steroid biosynthesis, and glycerophospholipid metabolism pathway in hepatocytes. The m6A methylation level decreased but the difference was not significant after METTL3 interference, and the expression levels of FABP4 and ATGL increased significantly (P < 0.05); the m6A methylation level significantly increased following METTL3 overexpression, and LPL and ATGL expression levels significantly decreased (P < 0.05), indicating that overexpression of METTL3 inhibited the expression of lipid deposition-related genes in a m6A-dependent manner. The m6A methylation level was significantly increased, ATGL expression was significantly decreased (P < 0.05), and LPL, FABP4, and Accα expression was not significantly changed following FTO interference (P > 0.05); the m6A methylation level was significantly decreased after FTO overexpression, and LPL, FABP4, and ATGL expression was significantly increased (P < 0.05), indicating that FTO overexpression increased the expression of lipid deposition-related genes in a m6A-dependent manner. Transcriptomic and metabolomic analyses showed that m6A methylation modification mainly regulated lipid metabolism through triglyceride metabolism, adipocytokine signaling, MAPK signaling, and fat digestion and absorption in hepatocytes. In the lipid deposition model of preadipocytes, the regulation of gene expression is the same as that in hepatocytes. CONCLUSIONS: METTL3 significantly inhibited the expression of lipid deposition-related genes, whereas FTO overexpression promoted lipid deposition. Our study provides a theoretical basis and reference for accurately regulating animal lipid deposition by mastering METTL3 and FTO genes to promote high-quality animal husbandry.

Fine-Scale Haplotype Mapping Reveals an Association of the FTO Gene with Osteoporosis and Fracture Risk in Postmenopausal Women.

INTRODUCTION: The Fat Mass and Obesity-Associated (FTO) gene encodes a demethylase, which modulates RNA N6-methyladenosine (m6A) and plays a regulatory role in adipocyte differentiation and the pathogenesis of human obesity. METHODS: To understand the potential role of FTO in osteoporosis (OP), we investigated five single nucleotide variations (SNVs) in intron 1 (rs8057044, rs8050136, rs9939609, rs62033406, and rs9930506) of the FTO gene, and a missense SNV i.e., rs3736228 (A1330V), located in exon 18 of the LRP5 gene, in a cohort of postmenopausal women (n = 188) from Central Europe. Genotyping was performed with an allele discrimination assay, while haplotypes were reconstructed in the population by PHASE 2.1. RESULTS: The rs9930506 was strongly associated with OP (p < 0.0035), which was supported by Bonferroni correction (p < 0.0175), and all SNVs located in the FTO gene were more strongly associated with severe OP with fragility fractures. Among seventeen haplotypes detected for the FTO gene, two haplotypes (H1 and H9) were frequent (frequency > 10%) and distributed in three main haplotypes pairs (H1/H1, H1/H9 and H9/H9, respectively). The pathogenic pair H1/H9 was associated with a leaner phenotype, increased fracture risk, and a lower bone mineral density (BMD), and carried the heterozygous GA of rs9930506, while the protective pair H9/H9 was associated with an increased obesity risk and carried AA alleles of rs9939609. CONCLUSIONS: Concordant associations with OP, an increased fracture risk, and a lower BMD at all skeletal sites indicate that the FTO gene is a promising candidate for OP, explaining the complex relationship with obesity and offering new perspectives for the study of the epigenetic regulation of bone metabolism.

Mechanism of USP18-Mediated NCOA4 m6A Modification Via Maintaining FTO Stability In Regulating Ferritinophagy-Mediated Ferroptosis in Cerebral Ischemia-Reperfusion Injury.

This study aimed to explore whether USP18 regulates cerebral ischemia-reperfusion (I/R) injury via fat mass and obesity-associated proteins (FTO)-mediated NCOA4. Middle cerebral artery occlusion (MCAO) models were established in mice, and PC-12 cells treated with oxygen-glucose deprivation and reperfusion (OGD/R) were used as in vitro models. The USP18 lentiviral vector was transfected into cells in vitro and MCAO mice to observe its effect on ferroptosis. The relationship between USP18 and FTO was assessed using Co-IP and western blot. The effect of FTO on NCOA4 m6A modification was also elucidated. Overexpression of USP18 in MCAO models decreased cerebral infarct size and attenuated pathological conditions in mouse brain tissues. Moreover, USP18 reduced iron content, MDA, ROS, and LDH release, increased GSH levels and cell viability in both MCAO models and OGD/R cells, and promoted LC3 expression and autophagy flux. In vitro experiments on neurons showed that USP18 maintained FTO stability. The presence of FTO-m6A-YTFDH1-NCOA4 was also verified in neurons. Both in vivo and in vitro experiments showed that FTO and NCOA4 abrogated the protective effects of USP18 against ferritinophagy-mediated ferroptosis. Notably, USP18 maintains FTO stability, contributing to the removal of NCOA4 m6A modification and the suppression of NCOA4 translation, which consequently inhibits ferritinophagy-mediated ferroptosis to attenuate cerebral I/R injury.

N6-methyladenosine demethylase FTO regulates neuronal oxidative stress via YTHDC1-ATF3 axis in arsenic-induced cognitive dysfunction.

Excessive exposure to metals in daily life has been proposed as an environmental risk factor for neurological disorders. Oxidative stress is an inevitable stage involved in the neurotoxic effects induced by metals, nevertheless, the underlying mechanisms are still unclear. In this study, we used arsenic as a representative environmental heavy metal to induce neuronal oxidative stress and demonstrated that both in vitro and in vivo exposure to arsenic significantly increased the level of N6-methyladenosine (m6A) by down-regulating its demethylase FTO. Importantly, the results obtained from FTO transgenic mice and FTO overexpressed/knockout cells indicated that FTO likely regulated neuronal oxidative stress by modulating activating transcription factor 3 (ATF3) in a m6A-dependent manner. We also identified the specific m6A reader protein, YTHDC1, which interacted with ATF3 and thereby affecting its regulatory effects on oxidative stress. To further explore potential intervention strategies, cerebral metabolomics was conducted and we newly identified myo-inositol as a metabolite that exhibited potential in protecting against arsenic-induced oxidative stress and cognitive dysfunction. Overall, these findings provide new insights into the importance of the FTO-ATF3 signaling axis in neuronal oxidative stress from an m6A perspective, and highlight a beneficial metabolite that can counteract the oxidative stress induced by arsenic.

Fat Mass- and Obesity-Associated Protein (FTO) Promotes the Proliferation of Goat Skeletal Muscle Satellite Cells by Stabilizing DAG1 mRNA in an IGF2BP1-Related m6A Manner.

Skeletal muscle development is spotlighted in mammals since it closely relates to animal health and economic benefits to the breeding industry. Researchers have successfully unveiled many regulatory factors and mechanisms involving myogenesis. However, the effect of N6-methyladenosine (m6A) modification, especially demethylase and its regulated genes, on muscle development remains to be further explored. Here, we found that the typical demethylase FTO (fat mass- and obesity-associated protein) was highly enriched in goats' longissimus dorsi (LD) muscles. In addition, the level of m6A modification on transcripts was negatively regulated by FTO during the proliferation of goat skeletal muscle satellite cells (MuSCs). Moreover, a deficiency of FTO in MuSCs significantly retarded their proliferation and promoted the expression of dystrophin-associated protein 1 (DAG1). m6A modifications of DAG1 mRNA were efficiently altered by FTO. Intriguingly, the results of DAG1 levels and its m6A enrichment from FB23-2 (FTO demethylase inhibitor)-treated cells were consistent with those of the FTO knockdown, indicating that the regulation of FTO on DAG1 depended on m6A modification. Further experiments showed that interfering FTO improved m6A modification at site DAG1-122, recognized by Insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) and consequently stabilized DAG1 transcripts. Our study suggests that FTO promotes the proliferation of MuSCs by regulating the expression of DAG1 through m6A modification. This will extend our knowledge of the m6A-related mechanism of skeletal muscle development in animals.

Recruitment of the m6A/m6Am demethylase FTO to target RNAs by the telomeric zinc finger protein ZBTB48.

BACKGROUND: N6-methyladenosine (m6A), the most abundant internal modification on eukaryotic mRNA, and N6, 2'-O-dimethyladenosine (m6Am), are epitranscriptomic marks that function in multiple aspects of posttranscriptional regulation. Fat mass and obesity-associated protein (FTO) can remove both m6A and m6Am; however, little is known about how FTO achieves its substrate selectivity. RESULTS: Here, we demonstrate that ZBTB48, a C2H2-zinc finger protein that functions in telomere maintenance, associates with FTO and binds both mRNA and the telomere-associated regulatory RNA TERRA to regulate the functional interactions of FTO with target transcripts. Specifically, depletion of ZBTB48 affects targeting of FTO to sites of m6A/m6Am modification, changes cellular m6A/m6Am levels and, consequently, alters decay rates of target RNAs. ZBTB48 ablation also accelerates growth of HCT-116 colorectal cancer cells and modulates FTO-dependent regulation of Metastasis-associated protein 1 (MTA1) transcripts by controlling the binding to MTA1 mRNA of the m6A reader IGF2BP2. CONCLUSIONS: Our findings thus uncover a previously unknown mechanism of posttranscriptional regulation in which ZBTB48 co-ordinates RNA-binding of the m6A/m6Am demethylase FTO to control expression of its target RNAs.

Association between FTO polymorphism and COVID-19 mortality among older adults: A population-based cohort study.

OBJECTIVES: COVID-19 caused a global pandemic with millions of deaths. Fat mass and obesity-associated gene (FTO) (alias m6A RNA demethylase) and its functional rs17817449 polymorphism are candidates to influence COVID-19-associated mortality since methylation status of viral nucleic acids is an important factor influencing viral viability. METHODS: We tested a population-based cohort of 5233 subjects (aged 63-87 years in 2020) where 70 persons died from COVID-19 and 394 from other causes during the pandemic period. RESULTS: The frequency of GG homozygotes was higher among those who died from COVID-19 (34%) than among survivors (19%) or deaths from other causes (20%), P <0.005. After multiple adjustments, GG homozygotes had a higher risk of death from COVID-19 with odds ratio = 2.01 (95% confidence interval; 1.19-3.41, P <0.01) compared with carriers of at least one T allele. The FTO polymorphism was not associated with mortality from other causes. CONCLUSIONS: Our results suggest that FTO variability is a significant predictor of COVID-19-associated mortality in Caucasians.

N6-methyladenosine demethyltransferase FTO alleviates sepsis by upregulating BNIP3 to induce mitophagy.

N6-methyladenosine (m6A) is known to be crucial in various biological processes, but its role in sepsis-induced circulatory and cardiac dysfunction is not well understood. Specifically, mitophagy, a specialized form of autophagy, is excessively activated during lipopolysaccharide (LPS)-induced myocardial injury. This study aimed to investigate the impact of LPS-induced endotoxemia on m6A-RNA methylation and its role in regulating mitophagy in sepsis-induced myocardial dysfunction. Our research demonstrated that FTO (fat mass and obesity-associated protein), an m6A demethylase, significantly affects abnormal m6A modification in the myocardium and cardiomyocytes following LPS treatment. In mice, cardiac dysfunction and cardiomyocyte apoptosis worsened after adeno-associated virus serotype 9 (AAV9)-mediated FTO knockdown. Further analyses to uncover the cellular mechanisms improving cardiac function showed that FTO reduced mitochondrial reactive oxygen species, restored both basal and maximal respiration, and preserved mitochondrial membrane potential. We revealed that FTO plays a critical role in activating mitophagy by targeting BNIP3. Additionally, the cardioprotective effects of AAV-FTO were significantly compromised by mdivi-1, a mitophagy inhibitor. Mechanistically, FTO interacted with BNIP3 transcripts and regulated their expression in an m6A-dependent manner. Following FTO silencing, BNIP3 transcripts with elevated m6A modification levels in their coding regions were bound by YTHDF2 (YT521-B homology m6A RNA-binding protein 2), leading to mRNA destabilization and decreased BNIP3 protein levels. These findings highlight the importance of FTO-dependent cardiac m6A methylation in regulating mitophagy and enhance our understanding of this critical interplay, which is essential for developing therapeutic strategies to protect cardiac mitochondrial function, alleviate cardiac dysfunction, and improve survival during sepsis.