Deep Sequencing Shows Multiple Oligouridylations Are Required for 3′ to 5′ Degradation of Histone mRNAs on Polyribosomes

• DOI: 10.1016/j.molcel.2014.02.027
• Publication Date: 2014-03-22T04:34:30Z
• Authors: Michael K. Slevin; Stacie Meaux; Joshua D. Welch; Rebecca Bigler; Paula L. Miliani ...; Wei Su; Robert E. Rhoads; Jan F. Prins; William F. Marzluff
• Keywords: 0301 basic medicine; RNA Stability; Molecular Sequence Data; S Phase; Histones; Jurkat Cells; Open Reading Frames; 03 medical and health sciences; Humans; Codon; Molecular Biology; 3' Untranslated Regions; Uridine; Gene Library; 0303 health sciences; Base Sequence; Exosome Multienzyme Ribonuclease Complex; Sequence Analysis, RNA; Gene Expression Regulation, Developmental; Cell Biology; Polyribosomes; Exoribonucleases; Nucleic Acid Conformation; HeLa Cells; Signal Transduction

Histone mRNAs are rapidly degraded when DNA replication is inhibited during S-phase with degradation initiating with oligouridylation of the stemloop at the 3′ end. We developed a customized RNA-Seq strategy to identify the 3′ termini of degradation intermediates of histone mRNAs. Using this strategy, we identified two types of oligouridylated degradation intermediates: RNAs ending at different sites of the 3′ side of the stemloop that resulted from initial degradation by 3′hExo and intermediates near the stop codon and within the coding region. Sequencing of polyribosomal histone mRNAs revealed that degradation initiates and proceeds 3′ to 5′ on translating mRNA and many intermediates are capped. Knockdown of the exosome-associated exonuclease Pml/Scl-100, but not the Dis3L2 exonuclease, slows histone mRNA degradation, consistent with 3′ to 5′ degradation by the exosome containing PM/Scl-100. Knockdown of No-go decay factors also slowed histone mRNA degradation, suggesting a role in removing ribosomes from partially degraded mRNAs.