Fig 2: YTHDF1 facilitates WWP1 expression in LPS + ATP-treated RAW264.7 cells.A The expression of WWP1 and four m6A readers in sepsis-related microarray GSE00159 (measurement data were expressed as mean ± standard deviation, and paired samples t test was applied for comparison between the control group and the sepsis group). B RT-qPCR to detect the expression of YTHDF1 in PBMCs of sepsis samples (n = 40) and healthy controls (n = 40). C The expression of YTHDF1 in PBMCs of mice with different treatment detected by RT-qPCR. D The expression of YTHDF1 determined by RT-qPCR. E The expression of YTHDF1 and WWP1 in cells with different treatment assessed by RT-qPCR. F Western blot assay to measure the protein expression of YTHDF1 and WWP1 in cells with different treatment. G The m6A modification status of WWP1 mRNA evaluated by meRIP. H, RIP-RT-qPCR to assess the binding of WWP1 mRNA to YTHDF1 protein. I Dual luciferase gene reporter assay to evaluate the binding of WWP1 and YTHDF1. There were 10 mice in each group. *p < 0.05; **p < 0.01; ***p < 0.001. ****p < 0.0001. ‘ns’ indicates no significant difference. Cell experiments were repeated three times.

Fig 3: The molecular mechanism of m6A reader protein YTHDF1 in sepsis.YTHDF1 upregulates WWP1 to promote NLRP3 ubiquitination, which inhibits caspase-1-dependent pyroptosis, thereby alleviating sepsis.

Fig 4: WWP1 is downregulated in sepsis.A The thermal map of differentially expressed genes in GSE100159 (12 normal control samples and 35 sepsis samples). B The intersection of UbiBrowser database and downregulated genes in GSE100159 dataset. C The expression of WWP1 in PBMCs collected from sepsis samples (n = 40) and healthy controls (n = 40) detected by RT-qPCR. D The survival rates of CLP-induced mice. E The pathological changes of lung, liver and kidney of CLP-induced mice observed by HE staining. F The expression of WWP1 in PBMCs of CLP-induced mice detected by RT-qPCR. There were ten mice in each group. **p < 0.01, ****p < 0.0001.

Fig 5: Upregulation of WWP1 by YTHDF1 inhibits activation of NLRP3 inflammasomes and caspase-1-mediated cleavage of GSDMD.A The expression of YTHDF1 and WWP1 in cells with different treatment assessed by RT-qPCR. B The expression of TNF-α and IL-1β determined by ELISA. C LDH level in cells with different treatment. D The activation of NLRP3 inflammasomes measured by Western blot assay. E Protein levels of GasD-N and GasD-FL detected by Western blot assay. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. ‘ns’ indicates no significant difference. Cell experiments were repeated 3 times.