Global analysis of mRNA decay intermediates in Saccharomyces cerevisiae

Abstract

The general pathways of eukaryotic mRNA decay occur via deadenylation followed by 3' to 5' degradation or decapping, although some endonuclease sites have been identified in metazoan mRNAs. To determine the role of endonucleases in mRNA degradation in Saccharomyces cerevisiae, we mapped 5' monophosphate ends on mRNAs in wild-type and dcp2 xrn1 yeast cells, wherein mRNA endonuclease cleavage products are stabilized. This led to three important observations. First, only few mRNAs that undergo low-level endonucleolytic cleavage were observed, suggesting that endonucleases are not a major contributor to yeast mRNA decay. Second, independent of known decapping enzymes, we observed low levels of 5' monophosphates on some mRNAs, suggesting that an unknown mechanism can generate 5' exposed ends, although for all substrates tested, Dcp2 was the primary decapping enzyme. Finally, we identified debranched lariat intermediates from intron-containing genes, demonstrating a significant discard pathway for mRNAs during the second step of pre-mRNA splicing, which is a potential step to regulate gene expression.