PEITC-mediated inhibition of mRNA translation is associated with both inhibition of mTORC1 and increased eIF2α phosphorylation in established cell lines and primary human leukemia cells.

Alison Yeomans,Mark J Coldwell,Breeze E Cavell,Freda K Stevenson,Andrew J Steele,Sarah Wilmore,Beatriz Valle-Argos,Sergey Krysov,Anne E Willis,Marina Sanchez Hidalgo,Francesco Forconi,Elizabeth Lemm,Elodie Leonard,Graham Packham

doi:10.18632/oncotarget.11655

Abstract

Increased mRNA translation drives carcinogenesis and is an attractive target for the development of new anti-cancer drugs. In this work, we investigated effects of phenethylisothiocyanate (PEITC), a phytochemical with chemopreventive and anti-cancer activity, on mRNA translation. PEITC rapidly inhibited global mRNA translation in human breast cancer-derived MCF7 cells and mouse embryonic fibroblasts (MEFs). In addition to the known inhibitory effects of PEITC on mTORC1 activity, we demonstrate that PEITC increased eIF2α phosphorylation. PEITC also increased formation of stress granules which are typically associated with eIF2α phosphorylation and accumulation of translationally stalled mRNAs. Analysis of genetically modified MEFs demonstrated that optimal inhibition of global mRNA translation by PEITC was dependent on eIF2α phosphorylation, but not mTORC1 inhibition. We extended this study into primary leukemic B cells derived from patients with chronic lymphocytic leukaemia (CLL). CLL cells were stimulated in vitro with anti-IgM to mimic binding of antigen, a major driver of this leukemia. In CLL cells, PEITC increased eIF2α phosphorylation, inhibited anti-IgM-induced mTORC1 activation and decreased both basal and anti-IgM-induced global mRNA translation. PEITC also inhibited transcription and translation of MYC mRNA and accumulation of the MYC oncoprotein, in anti-IgM-stimulated cells. Moreover, treatment of CLL cells with PEITC and the BTK kinase inhibitor ibrutinib decreased anti-IgM-induced translation and induced cell death to a greater extent than either agent alone. Therefore, PEITC can inhibit both global and mRNA specific translation (including MYC) via effects on multiple regulatory pathways. Inhibition of mRNA translation may contribute to the chemopreventive and anti-cancer effects of PEITC.

Highlights

Phenethyl isothiocyanate (PEITC) is a naturally occurring phytochemical that has received extensive interest for its anti-cancer and chemopreventive activities. [1] PEITC induces cell cycle arrest and apoptosis, and inhibits metastasis, proliferation and angiogenesis in various malignant cell types and, has inhibitory effects on multiple cancer hallmarks
We previously showed that the phytochemical PEITC inhibited mTORC1 activity and downstream HIF1A mRNA translation [4]
We demonstrated that PEITC rapidly inhibited global mRNA translation and used genetically-manipulated mouse embryonic fibroblasts (MEFs) to define the role of mTORC1 inhibition and eIF2α phosphorylation in this response

Summary

Introduction

Phenethyl isothiocyanate (PEITC) is a naturally occurring phytochemical that has received extensive interest for its anti-cancer and chemopreventive activities. [1] PEITC induces cell cycle arrest and apoptosis, and inhibits metastasis, proliferation and angiogenesis in various malignant cell types and, has inhibitory effects on multiple cancer hallmarks. Recent findings have indicated that PEITC inhibits mRNA translation [2,3,4]. MRNA translation has a high energy requirement and is subject to tight regulation, especially at the level of initiation. Translation initiation is mediated by eukaryotic initiation factors (eIF) that recruit the two subunits of the ribosome to the correct mRNA start codon and ensure delivery of the initiator methionine-loaded tRNA [6]. Additional eIFs are recruited to modulate translation initiation rate and stringency of start codon selection. EIF2B acts as a guanine nucleotide exchange factor for eIF2 (a trimer comprising α, β and γ subunits) and catalyzes the exchange of GDP to GTP that is required to form an active ternary complex after each round of translation initiation

Methods

Results

Conclusion