Extensive 5\u2032-surveillance guards against non-canonical NAD-caps of nuclear mRNAs in yeast

Yaqing Zhang,Hans Hombauer,Tobias Schmidt,Andres Jäschke,Vladimir Benes,David Ibberson,Gabriele Nübel,Daniel Kirrmaier,David Kuster,Michael Knop

doi:10.1038/s41467-020-19326-3

Yaqing Zhang, Hans Hombauer + Show 8 more

Open Access

https://doi.org/10.1038/s41467-020-19326-3

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Abstract

The ubiquitous redox coenzyme nicotinamide adenine dinucleotide (NAD) acts as a non-canonical cap structure on prokaryotic and eukaryotic ribonucleic acids. Here we find that in budding yeast, NAD-RNAs are abundant (>1400 species), short (<170 nt), and mostly correspond to mRNA 5′-ends. The modification percentage of transcripts is low (<5%). NAD incorporation occurs mainly during transcription initiation by RNA polymerase II, which uses distinct promoters with a YAAG core motif for this purpose. Most NAD-RNAs are 3′-truncated. At least three decapping enzymes, Rai1, Dxo1, and Npy1, guard against NAD-RNA at different cellular locations, targeting overlapping transcript populations. NAD-mRNAs are not translatable in vitro. Our work indicates that in budding yeast, most of the NAD incorporation into RNA seems to be disadvantageous to the cell, which has evolved a diverse surveillance machinery to prematurely terminate, decap and reject NAD-RNAs.

Highlights

The ubiquitous redox coenzyme nicotinamide adenine dinucleotide (NAD) acts as a noncanonical cap structure on prokaryotic and eukaryotic ribonucleic acids
As the protocol used in this work excluded the small RNA fraction, which had been rich in NAD-RNAs in prokaryotes[4], we address here the whole landscape of NAD transcripts in yeast using the original NAD captureSeq protocol[11]
Reverse transcription (RT), and PCR amplification, amplicons with sizes between 150 and 300 bp were selected by gel electrophoresis; this library represented mostly RNA species with sizes between 20 and 170 nt present in the original sample

Summary

Introduction

The ubiquitous redox coenzyme nicotinamide adenine dinucleotide (NAD) acts as a noncanonical cap structure on prokaryotic and eukaryotic ribonucleic acids. Our work indicates that in budding yeast, most of the NAD incorporation into RNA seems to be disadvantageous to the cell, which has evolved a diverse surveillance machinery to prematurely terminate, decap and reject NAD-RNAs. 1234567890():,; In eukaryotes, the 5′-terminus of messenger RNAs is protected by a m7-guanosine (m7G) cap[1], which modulates pre-mRNA splicing, polyadenylation, nuclear exit, and translation initiation[2]. The cap is hydrolyzed by various decapping enzymes[3], thereby triggering RNA degradation Another type of 5′cap structure was discovered, both in prokaryotes[4,5,6] and eukaryotes[7,8,9,10], which is derived from the ubiquitous redox coenzyme NAD. We speculate that the NAD modification is in most cases undesirable to the cell, which first disfavors the synthesis of full-length NAD-RNAs, decaps them rapidly using a multi-tiered machinery localized in different compartments and—even if they reach full length and escape decapping— rejects them from ribosomes

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