Targeting ATF4-dependent pro-survival autophagy to synergize glutaminolysis inhibition

作者：Shuting Han, Liyuan Zhu, Yiran Zhu, Yuan Meng, Jiaqiu Li, Ping Song, Neelum Aziz Yousafzai, Lifeng Feng, Miaoqin Chen, Yanmei Wang, Hongchuan Jin, Xian Wang · 发表于：Theranostics · 年份：2021 · DOI：10.7150/thno.60028 · 被引用次数：88 · 研究领域：Autophagy in Disease and Therapy、RNA modifications and cancer、Epigenetics and DNA Methylation

As glutamine plays a central role in cancer metabolism, inhibition of glutaminolysis has become an ideal anticancer therapeutic target. However, glutaminolysis inhibition leads to activation of autophagy, which compromises its antitumor effect. Hence, we investigated the mechanism underlying glutaminolysis inhibition-induced pro-survival autophagy. Methods: High-throughput sequencing was performed on colorectal cancer (CRC) cells before and after glutaminolysis inhibition to identify differentially expressed genes. Activating transcription factor 4 (ATF4) pathway enrichment in glutaminolysis inhibited cells was identified through gene set enrichment analysis. ATF4 expression was assessed by quantitative real-time PCR (qRT-PCR) and western blotting. The function of ATF4 on mechanistic target of rapamycin (mTOR) regulation was assessed by western blotting. Luciferase reporter assays and chromatin immunoprecipitation were used to confirm the regulation of DNA damage inducible transcript 4 (DDIT4) by ATF4. mRNA half-life assays, RNA immunoprecipitation, qRT-PCR and western blotting were performed to determine the relationship between FTO alpha-ketoglutarate dependent dioxygenase (FTO), YTH N 6 -methyladenosine RNA binding protein 2 (YTHDF2), and ATF4. ATF4 regulation of pro-survival autophagy was measured by tandem monomeric red fluorescent protein-green fluorescent protein fluorescence microscopy. Finally, the synergistic effect of autophagy and glutaminolysis inhibition was ana...