EGR2-mediated regulation of mA reader IGF2BP proteins drive RCC tumorigenesis and metastasis via enhancing S1PR3 mRNA stabilization.

Emerging discoveries of dynamic and reversible N6-methyladenosine (mA) modification on RNA in mammals have revealed the key roles of the modification in human tumorigenesis. As known mA readers, insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs) are upregulated in most cancers and mediates the enhancement of mA-modified mRNAs stability. However, the mechanisms of IGF2BPs in renal cell cancer (RCC) still remain unclear. Bioinformatic analysis and RT-qPCR were performed to evaluate the expression of IGF2BPs and mA writer Wilms tumor 1-associating protein (WTAP) in RCC samples and its correlation with patient prognosis. In vitro, in vivo biological assays were performed to investigate the functions of IGF2BPs and WTAP in RCC. Chromatin immunoprecipitation-qPCR (ChIP-qPCR) combined with bioinformatics analysis and following western blot assay, dual-luciferase reporter assays were performed to validate the regulatory relationships between transcription factor (TF) early growth response 2 (EGR2) and potential target genes IGF2BPs. RNA sequencing (RNA-seq), methylated RNA immunoprecipitation-qPCR (MERIP-qPCR), RIP-qPCR, mA dot blot, and dual-luciferase reporter assays combined with bioinformatics analysis were employed to screen and validate the direct targets of IGF2BPs and WTAP. Here, we showed that early growth response 2 (EGR2) transcription factor could increase IGF2BPs expression in RCC. IGF2BPs in turn regulated sphingosine-1-phosphate receptor 3 (S1PR3) expression in an mA-dependent manner by enhancing the stability of S1PR3 mRNA. They also promoted kidney tumorigenesis via PI3K/AKT pathway. Furthermore, IGF2BPs and WTAP upregulation predicted poor overall survival in RCC. Our studies showed that the EGR2/IGF2BPs regulatory axis and mA-dependent regulation of S1PR3-driven RCC tumorigenesis, which enrich the mA-modulated regulatory network in renal cell cancer. Together, our findings provide new evidence for the role of N6-methyladenosine modification in RCC.