ALYREF drives cancer cell proliferation through an ALYREF-MYC positive feedback loop in glioblastoma

Abstract

Background: While RNA-binding proteins (RBPs) are known to affect RNA homeostasis during cancer cell initiation and development, their characteristics and biological function in glioblastoma (GBM) remain unclear. Methods: Differences in RBP expression were explored by differential analysis of The Cancer Genome Atlas-GBM and Genotype-Tissue Expression (GTEx) datasets. Real-time PCR was conducted to verify the expressional levels of Aly/REF export factor (ALYREF) in normal brain and GBM tissues. Proliferative assays were performed to investigate molecular functions of ALYREF in GBM cells in vitro and in vivo. Real-time PCR and Kyoto Encyclopedia of Genes and Genomes (KEGG) were performed to analyze the ALYREF downstream signaling pathways. A chromatin immunoprecipitation (ChIP) assay was performed to identify key transcriptional factors that regulate ALYREF expression at RNA level. UV crosslinking, immunoprecipitation (CLIP) and RNA stability assays were conducted to reveal the bound RNAs and their stability regulated by ALYREF. Results: The results showed that ALYREF is frequently increased in GBM tissues, and its mRNA expression is regulated by the MYC proto-oncogene, bHLH transcription factor (MYC). Inhibition of ALYREF expression decreased GBM cell proliferative ability in vitro and tumor formation in vivo. KEGG analysis revealed that high ALYREF expression in GBM tissues was enriched in the upregulation of oncogenic pathways such as the Wnt/β-catenin signaling pathway. The CLIP assay showed that ALYREF drives GBM carcinogenesis by binding to and stabilizing MYC mRNAs. Overexpression of MYC restored the oncogenic property of ALYREF-deficient GBM cells. Conclusion: Our data showed that ALYREF is regulated by MYC at the transcriptional level. ALYREF drives GBM cell proliferation by activating the Wnt/β-catenin signaling pathway and stabilizing MYC mRNA, suggesting that an ALYREF-MYC positive feedback loop might be a potential therapeutic target for treating GBM patients.