The 3'UTR of the Pseudogene CYP4Z2P Promotes Tumor Angiogenesis in Breast Cancer by Acting As a CeRNA for CYP4Z1

Pseudogenes are now known to regulate their protein-coding counterparts. Additionally, disturbances of 3'UTRs could increase the risk of cancer susceptibility by acting as modulators of gene expression. The aim of this study was to investigate the roles of the pseudogene CYP4Z2P-3'UTR and functional gene CYP4Z1-3'UTR in breast cancer angiogenesis process. The levels of CYP4Z2P- and CYP4Z1-3'UTR and miRNA of interests were measured in 22 cancerous tissues paired with non-cancerous samples by qRT-PCR. The effects of CYP4Z2P- and CYP4Z1-3'UTR were studied by overexpression and RNA interference approaches in vitro and ex vivo. Insights of the mechanism of competitive endogenous RNAs were gained from bioinformatic analysis, luciferase assays, and western blot. The positive CYP4Z2P/CYP4Z1 interaction and negative interaction between predicted miRNAs and CYP4Z2P or CYP4Z1 were identified via qRT-PCR assay and bivariate correlation analysis. CYP4Z2P- and CYP4Z1-3'UTR share several miRNA-binding sites, including miR-211, miR-125a-3p, miR-197, miR-1226, and miR-204. The CYP4Z2P- and CYP4Z1-3'UTRs arrest the interference caused by of these miRNAs, resulting in increased translation of CYP4Z1. Moreover, ectopic expression of the CYP4Z2P- and CYP4Z1-3'UTRs exhibit tumor angiogenesis-promoting properties in breast cancer collectively by inducing the phosphorylation of ERK1/2 and PI3K/Akt. Co-transfection with Dicer siRNA reversed the CYP4Z2P 3'UTR-mediated changes. Additionally, PI3K or ERK inhibitors reversed CYP4Z2P- and CYP4Z1-3'UTR-mediated changes in VEGF-A expression. Increased CYP4Z2P- and CYP4Z1-3'UTR expression promotes tumor angiogenesis in breast cancer partly via miRNA-dependent activation of PI3K/Akt and ERK1/2. The CYP4Z2P- and CYP4Z1-3'UTRs could thus be used as combinatorial miRNA inhibitors.