Figure 6.

TFEB binding ABCA2 promoter to regulate its expression to involve PCa cell invasion and migration. (A) The binding motif of TFEB were provide from website. (B) The potential binding site of ABCA2 promoter. Mismatch rate is less than 1%. (C) Validation of the DNA fragment pulled down with TFEB chip-level antibody by qRT-PCR. DNA fragment were obtained from CUT&RUN assay and purified by DNA extraction kit. Rb IgG as a negative control. Anti-TFEB as an experimental group. (D) The DNA fragment product from qRT-PCR was validated by nucleic acid electrophoresis. The length of input, P1 and P2 mainly between 60 to 120 bp. (E, F) Transwell assay showed silenced ABCA2 expression inhibited PCa cell invasion. Cancer cells were stained after 24h. (G, H) Woundhealing assay showed silenced ABCA2 expression inhibited PCa cell migration after 48h culture. (I, J) Validation of ABCA2 LAMP1, MMP9, and MMP2 protein expression by western-blot after ABCA2 gene silenced. Quantitative analysis of the western-blot from (I). (K, L) DU145-vector and DU145-TFEB cell line were silenced ABCA2 for 72h and then treated with LysoTracker Red DND-99 (50 nM) for 45 min. Note: Statistical analysis was from three independent experiments and is presented as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001 compared with control group.