3′ Terminal oligo U-tract-mediated stimulation of decapping

Abstract

Decapping is a critical step in the control of gene expression and is regulated by both positive and negative trans factors. Less is known about cis elements that promote decapping. In plants, following microRNA (miRNA)-directed cleavage of an mRNA, a uridine tract can be added onto the exposed 39 end of the resulting 59 fragment, which can promote 59 end decay. We now demonstrate that in mammalian cell extract, addition of five uridine residues to the 39 end of an RNA (U 5 ) promotes decapping relative to an RNA lacking the uridines (U 0 ). Although the decapping stimulation observed in extract required hDcp2, recombinant hDcp2 was unable to support differential decapping of the U 0 and U 5 RNAs, indicating that the stimulation was likely due to an indirect recruitment of hDcp2 to the RNA. Consistent with the promotion of 59 end decapping by the uridine tract, affinity purification with the U 5 RNA revealed the presence of a decapping subcomplex at least consisting of hDcp2, Dcp1a, Edc4, LSm1, and LSm4 that were specifically bound to the U 5 RNA but not the U 0 RNA. In addition to promoting decapping, the U-tract stabilized the 39 end of the RNA by preventing 39 to 59 exonucleolytic decay to ensure 59 end directional degradation. These data suggest that following post-transcriptional oligo uridylation of an mRNA or mRNA fragment, the U-tract has the capacity to specifically stimulate 59 end decapping to expedite mRNA decay.

Methylation protects microRNAs from an AGO1-associated activity that uridylates 5′ RNA fragments generated by AGO1 cleavage

“…3 and 6). In fact, oligouridylation could prevent RNA from 3′ to 5′ degradation in vitro (36). However, we cannot rule out the possibility that 3′-to-5′ degradation activities triggered by uridylation are highly progressive, such that no or few 3′ truncation intermediates are accumulated in vivo.…”

“…S3). In humans and yeast, uridylation has been shown to induce decapping of some RNAs, followed by degradation (36)(37)(38). The ratio of uridylated MYB33-5′ in the uncapped population is higher than that in the capped population in Ler, suggesting that uridylation may also have a role in stimulating decapping.…”

A triple helix stabilizes the 3′ ends of long noncoding RNAs that lack poly(A) tails

“…1), microRNA precursors (Heo et al 2009), mature microRNAs (Li et al 2005), and transcription start site-associated RNAs (Choi et al 2012). Although there is currently less evidence for U-tails on long transcripts in vivo, uridylation can promote mRNA decapping (Song and Kiledjian 2007;Rissland and Norbury 2009), and U-tails have been observed on the products of microRNA-directed cleavage (Shen and Goodman 2004). Interestingly, histone mRNAs are subjected to uridylation and degradation following the completion of DNA synthesis (Mullen and Marzluff 2008).…”

Lsm1-7-Pat1 complex: A link between 3’ and 5’-ends in mRNA decay?

“…50 Second, in vitro mRNA decay assays carried out with mammalian cell extracts reveal that 3'-U-tracts can stimulate decapping of RNAs and that such stimulation is dependent on the presence of Lsm1 protein in the cell extract. 52 Moreover, several decapping factors including Lsm1 and Lsm4 proteins associate selectively to RNAs carrying such 3'-U-tracts when such RNAs are incubated with cell extracts. 52 These observations suggest that by virtue of its RNA binding preferences, the Lsm1-7-Pat1 complex could mediate the stimulation of mRNA decay (via the 5' to 3' pathway) by not only 3'-oligo(A) tails of mRNAs but also U-tracts near the 3'-ends of mRNAs.…”

“…52 Moreover, several decapping factors including Lsm1 and Lsm4 proteins associate selectively to RNAs carrying such 3'-U-tracts when such RNAs are incubated with cell extracts. 52 These observations suggest that by virtue of its RNA binding preferences, the Lsm1-7-Pat1 complex could mediate the stimulation of mRNA decay (via the 5' to 3' pathway) by not only 3'-oligo(A) tails of mRNAs but also U-tracts near the 3'-ends of mRNAs.…”