Fig. 6. METTL1 deficiency elicits SASP through ribotoxic stress response and integrated stress response.

a Ubiquitination of RPS20 in isolated ribosomes from indicated samples was detected using western blotting. ANS-treated IMR90 cells served as the positive control. Arrow-pointed bands indicated ubiquitination of the RPS20 protein. b ISR and RSR response-relevant proteins, were detected by western blotting in proliferating/senescent IMR90 cells(b), young/aged liver tissues, n = 5 mice per group(2 female and 3 male) (c), METTL1 KD/control cells(d), liver tissues from Mettl1 KO and control, n = 5 mice per group(3 female and 2 male) (e), and METTL1 and mutated AFPA mutant transduced IMR90 cells (f) were detected using western blotting. g Phosphorylated eIF2α and ATF4 proteins were detected in senescent cells treated with various inhibitors using western blotting. h Ribotoxic stress response proteins were examined upon the inhibition of ZAKα by delivering siRNAs into METTL1 KD cells. i SASP factors were measured by RT-qPCR in METTL1 knockdown cells with or without inhibitors, as indicated. j The IL-6 levels in Mettl1 knockout (KO) mice with or without inhibitor and control mice were detected using ELISA (n = 8 per group, 4 female and 4 male). k The impact of GCN2, p38, and JNK inhibitors on Mettl1-mediated senescence status was assessed through SA-β-gal staining. Scale bar = 100 μm. All in vitro assays were biologically repeated three times. All WB results were performed three times, the representative result was shown, the protein intensity was quantified with Image J (a, c, e); All data were presented as the mean ± SEM. Two-tailed unpaired t test (a), one-way ANOVA with Bonferroni’s multiple comparisons test (c, e, i–k). *p  <  0.05, **p  <  0.01, ***p  <  0.001. Source data are provided as a Source Data file.