Fig. 4

Ethionine changes m6A modification through abnormal expression of m6A methylase and demethylase. A RT-qPCR was used to detect mRNA level of N6-methyladenosine RNA methylase and demethylase in normal and NTDs embryo brain tissue at E10.5. *p < 0.05, **p < 0.01 and ***p < 0.001 vs control. B ELISA was used to detect m6A levels in normal and NTDs embryo brain tissue at E10.5. ***p < 0.001 and ****p < 0.0001 vs control. C Flow cytometry was used to detect the effect of SiRNA transfection, and SiNc was used as a negative control. D Mettl3 gene expression as assessed by RT-qPCR analysis in HT-22 cells. Mean ± SEM *p < 0.05, **p < 0.01 or ***p < 0.001 vs. The β-actin gene was used as a control. E, F Western blot analysis detected the METTL3 protein level in the SiMettl3 and SiNc groups was evaluated, and the expression of β-Tubulin was used as the loading control. G, H Western blot analysis of the protein levels of ALKBH5 in the HT-22 cells treated with control OE and Alkbh5 OE. β-Tubulin was used as a loading control. Bar graphs for protein abundance were quantitative data from three independent experiments. (I) Alkbh5 gene expression as assessed by RT-qPCR analysis in HT-22 cells treated with control OE and Alkbh5 OE. Mean ± SEM ***p < 0.001 vs. The β-actin gene was used as a control. (J) ELISA was used to detect m6A levels in SiMettl3 and SiNc cells. ****p < 0.0001 vs control. K ELISA was used to detect m6A levels in control OE and Alkbh5 OE cells. **p < 0.01 vs control