RNF219 attenuates global mRNA decay through inhibition of CCR4-NOT complex-mediated deadenylation

Fabian Poetz,Doris Lindner,Johanna Schott,Yevgen Levdansky,Vera Magg,Svetlana Lebedeva,Joshua Corbo,Georg Stoecklin,Eugene Valkov,Alexander Spiegelhalter,Jörg Schweiggert

doi:10.1038/s41467-021-27471-6

Abstract

The CCR4-NOT complex acts as a central player in the control of mRNA turnover and mediates accelerated mRNA degradation upon HDAC inhibition. Here, we explored acetylation-induced changes in the composition of the CCR4-NOT complex by purification of the endogenously tagged scaffold subunit NOT1 and identified RNF219 as an acetylation-regulated cofactor. We demonstrate that RNF219 is an active RING-type E3 ligase which stably associates with CCR4-NOT via NOT9 through a short linear motif (SLiM) embedded within the C-terminal low-complexity region of RNF219. By using a reconstituted six-subunit human CCR4-NOT complex, we demonstrate that RNF219 inhibits deadenylation through the direct interaction of the α-helical SLiM with the NOT9 module. Transcriptome-wide mRNA half-life measurements reveal that RNF219 attenuates global mRNA turnover in cells, with differential requirement of its RING domain. Our results establish RNF219 as an inhibitor of CCR4-NOT-mediated deadenylation, whose loss upon HDAC inhibition contributes to accelerated mRNA turnover.

Highlights

The carbon catabolite repression 4 (CCR4)-negative on TATA-less (NOT) complex acts as a central player in the control of mRNA turnover and mediates accelerated mRNA degradation upon HDAC inhibition
We previously discovered that protein acetylation has a profound impact on global mRNA turnover[53], whereby inhibition of HDAC1 and HDAC2 induces widespread degradation of bulk poly(A) RNA in mammalian and Drosophila melanogaster (Dm) cells
Individual HeLa clones were tested for FST-NOT1 expression, and Sanger sequencing of the targeted locus in clone #47, which was used for all subsequent experiments, confirmed in-frame integration of the FST sequence downstream of the start codon in one CNOT1 allele (Supplementary Fig. 1)

Summary

Introduction

The CCR4-NOT complex acts as a central player in the control of mRNA turnover and mediates accelerated mRNA degradation upon HDAC inhibition. Our results establish RNF219 as an inhibitor of CCR4-NOT-mediated deadenylation, whose loss upon HDAC inhibition contributes to accelerated mRNA turnover. Regulation of mRNA stability represents a key step in the control of post-transcriptional gene expression and shapes the kinetics of critical cellular processes such as cell proliferation, differentiation, immunity, and development[1,2,3,4,5,6,7]. (CCR4)-negative on TATA-less (NOT) complex functions as the principal deadenylase responsible for processive poly(A) tail shortening in eukaryotic cells and is essential for normal rates of mRNA turnover[17,18,19,20,21].

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Results

Conclusion