GSE175417
Revealing new molecular pathways for cancer cell fitness through a genetic screen of the cancer translatome
The major cap binding protein eIF4E, an ancient protein required for translation of all eukaryotic genomes, is a surprising yet potent oncogenic driver. The genetic interactions that maintain the oncogenic activity of this key translation factor remain unknown. Here we carried out a genome-wide CRISPRi screen wherein we identified over 600 genetic interactions that sustain eIF4E oncogenic activity. Our data show that eIF4E controls the translation of Tfeb, a key executer of the autophagy response. This autophagy survival response is triggered by mitochondrial proteotoxic stress, which allows cancer cell survival. Our screen also reveals a functional interaction between eIF4E and a single anti-apoptotic factor, Bcl-xL, in tumor growth. Furthermore, we show that eIF4E and the exon-junction complex (EJC), involved in many steps of RNA metabolism, interact to control the migratory properties of cancer cells. Overall, we have uncovered several cancer-specific vulnerabilities that provide unprecedented resolution of the cancer translatome. Overall design: eIF4E+/+ and eIF4E+/- MEFs expressing scramble, Magoh, eIF4A3 and Rbm8a sgRNAs were induced with Dox for 48 hours in triplicates, total RNA was isolated by Trizol and ribosomal RNA was removed by Ribo-Zero (Illumina). RNA seq libraries were prepared by ScriptSeq v2 RNA-seq library preparation kit (Epicenter) following the manufacturer's protocol and sequenced as paired-end 100 bp on HiSeq 4000 by the Center for Advance Technology core at UCSF. RNA-seq reads were aligned to mouse GRCm38 genome by using HISAT v2.0.5 (Kim et al., 2015)with default settings. Gene read counts and expression values (measured by Transcripts Per Kilobase Million, TPM) were further calculated by RSEM v1.3.0 (Li and Dewey, 2011) with default settings. Differential expression analyses were conducted with DESeq2 (Love et al., 2014) R package, where only genes with an average TPM over 0.5 were involved.