CNOT3 suppression promotes necroptosis by stabilizing mRNAs for cell death-inducing proteins

Toru Suzuki; Chisato Kikuguchi; Sahil Sharma; Toshio Sasaki; Miho Tokumasu; Shungo Adachi; Tohru Natsume; Yumi Kanegae; Tadashi Yamamoto

doi:10.1038/srep14779

CNOT3 suppression promotes necroptosis by stabilizing mRNAs for cell death-inducing proteins

Sci Rep. 2015 Oct 6:5:14779. doi: 10.1038/srep14779.

Authors

Toru Suzuki¹, Chisato Kikuguchi¹, Sahil Sharma¹, Toshio Sasaki², Miho Tokumasu¹, Shungo Adachi³, Tohru Natsume³, Yumi Kanegae⁴, Tadashi Yamamoto¹

Affiliations

¹ Cell Signal Unit, Okinawa Institute of Science and Technology, 1919-1 Onna-son, Okinawa 904-0495, Japan.
² Imaging and Instrumental Analysis Section, Okinawa Institute of Science and Technology Graduate University, 1919-1 Onna-son, Okinawa 904-0495, Japan.
³ Molecular Profiling Research Center for Drug Discovery, National Institute of Advanced Industrial Science and Technology, Tokyo 135-0064, Japan.
⁴ Laboratory of Molecular Genetics, Institute of Medical Science, University of Tokyo, Tokyo 108-8639, Japan.

Abstract

The CCR4-NOT complex is conserved in eukaryotes and is involved in mRNA metabolism, though its molecular physiological roles remain to be established. We show here that CNOT3-depleted mouse embryonic fibroblasts (MEFs) undergo cell death. Levels of other complex subunits are decreased in CNOT3-depleted MEFs. The death phenotype is rescued by introduction of wild-type (WT), but not mutated CNOT3, and is not suppressed by the pan-caspase inhibitor, zVAD-fluoromethylketone. Gene expression profiling reveals that mRNAs encoding cell death-related proteins, including receptor-interacting protein kinase 1 (RIPK1) and RIPK3, are stabilized in CNOT3-depleted MEFs. Some of these mRNAs bind to CNOT3, and in the absence of CNOT3 their poly(A) tails are elongated. Inhibition of RIPK1-RIPK3 signaling by a short-hairpin RNA or a necroptosis inhibitor, necrostatin-1, confers viability upon CNOT3-depleted MEFs. Therefore, we conclude that CNOT3 targets specific mRNAs to prevent cells from being disposed to necroptotic death.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Amino Acid Chloromethyl Ketones / pharmacology
Animals
Caspase Inhibitors / pharmacology
Cell Death / drug effects
Cell Survival / drug effects
Cysteine Proteinase Inhibitors / pharmacology
Embryo, Mammalian
Fibroblasts / cytology
Fibroblasts / drug effects
Fibroblasts / metabolism*
Gene Expression Regulation
Imidazoles / pharmacology
Indoles / pharmacology
Leupeptins / pharmacology
Mice
Primary Cell Culture
Protein Binding
RNA Stability
RNA, Messenger / genetics*
RNA, Messenger / metabolism
RNA, Small Interfering / genetics
RNA, Small Interfering / metabolism
Receptor-Interacting Protein Serine-Threonine Kinases / antagonists & inhibitors
Receptor-Interacting Protein Serine-Threonine Kinases / genetics*
Receptor-Interacting Protein Serine-Threonine Kinases / metabolism
Signal Transduction
Transcription Factors / genetics*
Transcription Factors / metabolism

Substances

Amino Acid Chloromethyl Ketones
CNOT3 protein, mouse
Caspase Inhibitors
Cysteine Proteinase Inhibitors
Imidazoles
Indoles
Leupeptins
RNA, Messenger
RNA, Small Interfering
Transcription Factors
benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone
necrostatin-1
Receptor-Interacting Protein Serine-Threonine Kinases
Ripk1 protein, mouse
Ripk3 protein, mouse
benzyloxycarbonylleucyl-leucyl-leucine aldehyde