Extensive 5’-Surveillance Guards Against Non-Canonical NAD-Caps of Nuclear mRNAs in Yeast

Summary 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 is low (<5%). NAD is incorporated during the initiation step 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 do not support translation in vitro . Our work indicates that in budding yeast, most of the NAD incorporation into RNA seems to be accidental and undesirable to the cell, which has evolved a diverse surveillance machinery to prematurely terminate, decap and reject NAD-RNAs. In Brief In budding yeast, most of the NAD incorporation into RNA seems to be accidental and undesirable to the cell, which has evolved a diverse surveillance machinery to prematurely terminate, decap and reject NAD-RNAs. Graphical Abstract Highlights Yeast cells have thousands of short NAD-RNAs related to the 5’-ends of mRNAs RNA polymerase II prefers a Y A AG promoter motif for NAD incorporation into RNA NAD-RNA is strongly guarded against by Rai1, Dxo1, and Npy1 decapping enzymes at different subcellular sites In vitro , NAD-mRNAs are rejected from translation