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ER-stress-induced transcriptional regulation increases protein synthesis leading to cell death

Abstract

Protein misfolding in the endoplasmic reticulum (ER) leads to cell death through PERK-mediated phosphorylation of eIF2α, although the mechanism is not understood. ChIP-seq and mRNA-seq of activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP), key transcription factors downstream of p-eIF2α, demonstrated that they interact to directly induce genes encoding protein synthesis and the unfolded protein response, but not apoptosis. Forced expression of ATF4 and CHOP increased protein synthesis and caused ATP depletion, oxidative stress and cell death. The increased protein synthesis and oxidative stress were necessary signals for cell death. We show that eIF2α-phosphorylation-attenuated protein synthesis, and not Atf4 mRNA translation, promotes cell survival. These results show that transcriptional induction through ATF4 and CHOP increases protein synthesis leading to oxidative stress and cell death. The findings suggest that limiting protein synthesis will be therapeutic for diseases caused by protein misfolding in the ER.

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Figure 1: ATF4 and CHOP bind to promoter regions of genes encoding protein synthesis and UPR functions.
Figure 2: ATF4 and CHOP upregulate expression of genes encoding protein synthesis and UPR functions.
Figure 3: ATF4 and CHOP interact to induce target genes involved in protein synthesis and the UPR.
Figure 4: ATF4 and CHOP increase protein synthesis leading to cell death.
Figure 5: ATF4 and CHOP increase oxidative stress and deplete ATP.
Figure 6: ATF4 and CHOP increase protein synthesis and oxidative stress in vivo.

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Acknowledgements

We thank the University of Michigan Sequencing Core for next-generation sequencing, D. Ron (University of Cambridge, UK) for recombinant retroviral vectors expressing GADD34 derivatives and GFP and Ratan (Cornell University, USA) for adenovirus expressing ATF4 and ATF4ΔRK. We are grateful to members of the R.J.K. laboratory for assistance, advice and stimulating discussions. This work was supported by a University of Michigan CCMB Pilot Grant (J.H.), NIH grants (HL057346, DK042394, DK088227, HL052173, DK093074 (R.J.K.)), (DK092062, DK094729 (M.S.K.)), (DK60596, DK53307 (M.H.)), and the National Research Foundation of Korea (NRF) grants 2010-0001199, 2011-0011433, 2012M3A9C3048686 (S.H.B.).

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J.H., S.H.B. and R.J.K. designed all experiments and performed most of them. J.H., Y-H.L and M.A.S. performed bioinformatic analysis and contributed to preparation of figures and tables. R.G. performed western blot and cell viability assays using Eif2 αA/A and Atf4−/− MEFs and GADD34 overexpression experiments. J.S. and M.S.K. generated and characterized the ATF4 antibody and performed co-immunoprecipitation and sequential ChIP experiments. C.L.Y., D.K. and M.H. measured in vivo protein synthesis using 2H2O. S.W. analysed protein synthesis and western blots. J.H., S.H.B. and R.J.K. prepared the manuscript.

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Correspondence to Randal J. Kaufman.

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Han, J., Back, S., Hur, J. et al. ER-stress-induced transcriptional regulation increases protein synthesis leading to cell death. Nat Cell Biol 15, 481–490 (2013). https://doi.org/10.1038/ncb2738

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