The E3 Ubiquitin Ligase ARIH1 Protects against Genotoxic Stress by Initiating a 4EHP-Mediated mRNA Translation Arrest

Abstract

bDNA damage response signaling is crucial for genome maintenance in all organisms and is corrupted in cancer. In an RNA interference (RNAi) screen for (de)ubiquitinases and sumoylases modulating the apoptotic response of embryonic stem (ES) cells to DNA damage, we identified the E3 ubiquitin ligase/ISGylase, ariadne homologue 1 (ARIH1). Silencing ARIH1 sensitized ES and cancer cells to genotoxic compounds and ionizing radiation, irrespective of their p53 or caspase-3 status. Expression of wild-type but not ubiquitinase-defective ARIH1 constructs prevented sensitization caused by ARIH1 knockdown. ARIH1 protein abundance increased after DNA damage through attenuation of proteasomal degradation that required ATM signaling. Accumulated ARIH1 associated with 4EHP, and in turn, this competitive inhibitor of the eukaryotic translation initiation factor 4E (eIF4E) underwent increased nondegradative ubiquitination upon DNA damage. Genotoxic stress led to an enrichment of ARIH1 in perinuclear, ribosome-containing regions and triggered 4EHP association with the mRNA 5= cap as well as mRNA translation arrest in an ARIH1-dependent manner. Finally, restoration of DNA damage-induced translation arrest in ARIH1-depleted cells by means of an eIF2 inhibitor was sufficient to reinstate resistance to genotoxic stress. These findings identify ARIH1 as a potent mediator of DNA damage-induced translation arrest that protects stem and cancer cells against genotoxic stress. DNA damage leads to acute toxicity and the accumulation of mutations and chromosomal instability, potentially resulting in malignant transformation (1, 2). To counteract these deleterious effects of DNA damage, the cell is equipped with a highly complex signaling response termed the DNA damage response (DDR). The DDR activates effector components involved in protective pathways, including DNA damage repair, cell cycle arrest, transcription regulation, chromatin remodeling, and cell death (1). The complex of DDR signaling pathways is crucial for the protection of the genome in all organisms. Moreover, understanding DDR signaling in the context of chemical or ionizing radiation-induced DNA damage is important to design improved strategies to combat therapy resistance. In tandem with phosphorylation-mediated signaling, which is largely executed by the phosphoinositol 3-kinase (PI3K)-like kinases ATM, ATR, and DNA-dependent protein kinase (DNA-PK), the checkpoint kinases Chk1 and Chk2, and members of the mitogen-activated protein kinase (MAPK) family (3, 4), protein modifications by ubiquitin and ubiquitin-like moieties are crucial at all levels of the DDR (5).The ubiquitination machinery can form various, differentially interpreted tags, including both degradative (K48-and K11-linked chains) and nondegradative (monoubiquitination and K63-linked chains) signals (6). Furthermore, a growing family of ubiquitin-like modifications, such as SUMO, Nedd8, and ISG15, has been identified, mostly providing nondegradative signals. Multiple enzymes are shared between the ubiquitinat...

Genome‐wide screening identifies novel genes and biological processes implicated in cisplatin resistance

“…ARIH1 has also been shown to protect against cisplatin-induced apoptosis by ubiquitinating EIF4E2, irrespective of the status of p53 or caspase-3 (22). The ubiquitinated EIF4E2 promotes translation arrest by competing with eukaryotic translation initiation factor 4E for binding to the mRNA cap (22). In agreement with the previous reports, we found that ARIH1 is overexpressed in A375 melanoma cells and contributes to cisplatin resistance in a p53-independent manner.…”

“…We searched for NF2-regulated factors that might be implicated in cisplatin resistance. We found that EIF4E2 (also called 4EHP), which represses global translation initiation by competing with eukaryotic translation initiation factor 4E for binding to the mRNA cap (21), and ARIH1, an E3 ubiquitin ligase that promotes binding of EIF4E2 to the mRNA cap by ubiquitinating EIF4E2 in response to DNA damage (22), were barely detectable in the Hermes 4C melanocyte and P1F/TERT fibroblast cells but were highly expressed in A375 melanoma cells (Fig. 7B).…”

“…ARIH1 has been shown recently to be widely overexpressed in cancer cells, notably in breast and lung adenocarcinomas, and to protect against cisplatin-induced cell death by promoting clearance of damaged mitochondria (mitophagy) (45). ARIH1 has also been shown to protect against cisplatin-induced apoptosis by ubiquitinating EIF4E2, irrespective of the status of p53 or caspase-3 (22). The ubiquitinated EIF4E2 promotes translation arrest by competing with eukaryotic translation initiation factor 4E for binding to the mRNA cap (22).…”

New frontiers in translational control of the cancer genome

“…One unexpected finding came when 4EHP (also known as eIF4E2), a cap-binding homologue of eIF4E best characterized for its role as a translational suppressor crucial for embryonic development and stem cell function 187–189 , was implicated in the hypoxic stress response. Although eIF4E is typically thought to be the primary cap-binding protein under hypoxic conditions 190 , 4EHP was recently identified as a central component of a hypoxia-induced translation initiation complex that includes oxygen-regulated hypoxia-inducible factor 2α (HIF2α) and the RBP RBM4 191 .…”

GIGYF1/2 proteins use auxiliary sequences to selectively bind to 4EHP and repress target mRNA expression

“…Furthermore, because 4EHP has a reduced affinity for the cap structure compared with eIF4E (Rom et al 1998;Zuberek et al 2007), it is possible that the auxiliary sequence-mediated dimerization observed in this study may have some as yet undefined functions (e.g., increasing local concentration of repressor complexes on the mRNA), but this hypothesis needs to be tested in future studies. The affinity of 4EHP for capped mRNAs may also be stimulated by post-translational modifications such as ISG15 modification (Okumura et al 2007) and monoubiquitinylation/ diubiquitinylation (von Stechow et al 2015), but whether these are synergistic with the GYF1/2 proteins is currently not known.…”