Fig. 4

The relationship between TFAP2C and STEAP3. (A) The correlation between TFAP2C and STEAP3 was analyzed using the GEPIA database. (B) The mRNA levels of TFAP2C in LUSC and adjacent normal tissues were determined using the TCGA database. (C) A total of 40 paired LUSC and adjacent normal lung tissues were collected for mRNA level analysis of TFAP2C. (D) The correlation between TFAP2C and STEAP3 was analyzed using our own collected clinical samples. (E) The protein levels of TFAP2C in LUSC and adjacent normal tissues were analyzed by immunohistochemistry. Scale bar represents 100 μm. (F) The Kaplan-Meier Plotter database was employed to compare overall survival (OS) between groups with disparate TFAP2C expression levels. (G) The SK-MES-1 cells were transfected with a recombinant plasmid carrying three different shRNAs targeting TFAP2C. The NCI-H520 cells were transfected with a recombinant plasmid carrying full-length CDS region of TFAP2C. The mRNA levels of TFAP2C were examined using real-time PCR. GAPDH was used as the internal control. (H) The protein levels of TFAP2C were quantified by western blot analysis. β-actin was employed as the internal control. (I) Following the knockdown and overexpression of TFAP2C, the cells were subjected to real-time PCR analysis of the mRNA levels of STEAP3. GAPDH was employed as the internal control. (J) The protein levels of STEAP3 were quantified by western blot analysis following the knockdown and overexpression of TFAP2C. β-actin was employed as the internal control. (K) The JASPAR database was used to predict the binding motif of TFAP2C. (L) The binding of transcription factor TFAP2C to the promoter of STEAP3 was examined by ChIP assay. (M) The regulatory relationship between TFAP2C and STEAP3 was analyzed using a dual-luciferase reporter assay. *P < 0.05 and **P < 0.01