Fig. 4.
YTHDF2-facilitated decay of YAP mRNA is mediated by AGO2 system and regulated by m6A modification. (a) The mRNA and protein levels of YTHDF2 were analyzed in paired tumor cancer tissues (Tumor) and adjacent normal tissues (Normal) coming from BinZhou medical university hospital (left panel, n = 10; right panel, n = 30). (b) The expression of YTHDF2 was analyzed by IHC assay in the different grades of lung cancer tissues. (c) Low expression of YTHDF2 is associated with worse prognosis for NSCLC patients (P = 0.021). (d-g) A549 cells were transfected with indicated genes of YTHDF2. (d) The expressions of YTHDF2 were analyzed by RT-PCR and western blot assays. (e) The cellular growth was analyzed by CCK8 assay. (f) The invasion and migration growths were analyzed by transwell assay. (g) The expressions of E-cadherin and Vimentin were analyzed by western blot assay. (h) The mRNA levels of YAP, CTGF and Cyr61 were detected in A549 cells by qPCR assay. (i) Co-IPs performed using lysates collected from A549 cells with immunoprecipitation by either YTHDF2 or AGO2 antibodies, respectively. (j-l) The mRNA level of YAP was analyzed by qPCR in Actinomycin D treated A549 cells with transfected with indicated genes. (m-q) A549 cell were transfected with indicated genes of YTHDF2 and YAP. (m) The cellular growth was analyzed by CCK8 assay. (n) The size and number of colons were analyzed by colony formation assay. (o) The cellular migration growth was analyzed by scratch assay. (p) The expressions of E-cadherin and Vimentin were analyzed by qPCR assay. (q) The relative of cleaved Caspas-3 (Caspas-3-Cl) was analyzed by western blot. Results were presented as mean ± SD of three independent experiments. *P < 0.05 or **P < 0.01 indicates a significant difference between the indicated groups. ns, not significant