Degradation of YRA1 Pre-mRNA in the Cytoplasm Requires Translational Repression, Multiple Modular Intronic Elements, Edc3p, and Mex67p
and Mex67p. PLoS Biol 8(4): e1000360. doi:10.1371/journal.pbio.1000360
Degradation of YRA1 Pre-mRNA in the Cytoplasm Requires Translational Repression, Multiple Modular Intronic Elements, Edc3p, and Mex67p
Shuyun Dong 0
Allan Jacobson 0
Feng He 0
Marv Wickens, University of Wisconsin, United States of America
0 Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School , Worcester, Massachusetts , United States of America
Intron-containing pre-mRNAs are normally retained and processed in the nucleus but are sometimes exported to the cytoplasm and degraded by the nonsense-mediated mRNA decay (NMD) pathway as a consequence of their inclusion of intronic in-frame termination codons. When shunted to the cytoplasm by autoregulated nuclear export, the introncontaining yeast YRA1 pre-mRNA evades NMD and is targeted by a cytoplasmic decay pathway mediated by the decapping activator Edc3p. Here, we have elucidated this transcript-specific decay mechanism, showing that Edc3p-mediated YRA1 pre-mRNA degradation occurs independently of translation and is controlled through five structurally distinct but functionally interdependent modular elements in the YRA1 intron. Two of these elements target the pre-mRNA as an Edc3p substrate and the other three mediate transcript-specific translational repression. Translational repression of YRA1 premRNA also requires the heterodimeric Mex67p/Mtr2p general mRNA export receptor, but not Edc3p, and serves to enhance Edc3p substrate specificity by inhibiting the susceptibility of this pre-mRNA to NMD. Collectively, our data indicate that YRA1 pre-mRNA degradation is a highly regulated process that proceeds through translational repression, substrate recognition by Edc3p, recruitment of the Dcp1p/Dcp2p decapping enzyme, and activation of decapping.
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Funding: This work was supported by a grant to AJ (R37 GM27757) from the National Institutes of Health. The funder had no role in study design, data collection
and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
mRNA degradation controls the level of gene expression and
ensures transcript quality control. In the yeast Saccharomyces
cerevisiae, most wild-type mRNAs are degraded by the general 59
to 39 or 39 to 59 decay pathways [1]. Functionally impaired
mRNAs are targeted for degradation by several
translationdependent mRNA surveillance mechanisms, including
nonsensemediated mRNA decay (NMD) for mRNAs containing premature
termination codons [2], non-stop decay (NSD) for mRNAs lacking
translation termination codons [3,4], and no-go decay (NGD) for
mRNAs stalled in translational elongation [5]. Transcript-specific
decay pathways have also been identified in several experimental
systems [6,7]. In each of these pathways, degradation of a
transcript is regulated by specific cis-acting elements and their
respective trans-regulatory RNA-binding factors. For example,
adenine/uridine-rich elements (AREs) have been found in the
39untranslated regions (39-UTRs) of diverse eukaryotic mRNAs [8],
and these elements by themselves, or through their interacting
proteins, can accelerate transcript-specific decay by recruitment of
the PARN and Ccr4p deadenylases [9,10], the exosome [1113],
or the Dcp1p/Dcp2p decapping enzyme [9]. Our recent
experiments, and those of Badis et al., have identified a yeast
cytoplasmic, transcript-specific decay pathway, mediated by the
decapping activator Edc3p, that principally targets only two
transcripts, RPS28B mRNA and intron-containing YRA1
premRNA [14,15].
Intron-containing pre-mRNAs are normally retained and
processed in the nucleus [16,17] but are sometimes exported to the
cytoplasm where their inclusion of intronic in-frame termination
codons targets these transcripts for degradation by the NMD
pathway [18,19]. However, the intron-containing YRA1 pre-mRNA
evades NMD and is degraded by the Edc3p-mediated decay
pathway [15]. Importantly, this Edc3p-mediated YRA1 pre-mRNA
decay is dependent on the presence of the YRA1 intron and appears
to require the function of the general mRNA export factor Mex67p
[15]. Here, we have dissected the intronic decay element and the
role of Mex67p in Edc3p-mediated YRA1 pre-mRNA decay. Our
experiments delineated five structurally distinct but functionally
interdependent cis-acting modules within the intron. Two modules
dictate Edc3p substrate specificity and are designated as Edc3p
responsive elements (EREs), whereas the other three modules,
designated as translational repression elements (TREs), inhibit the
translation of YRA1 pre-mRNA. This translational repression
requires Mex67p and Mtr2p, but not Edc3p, and prevents YRA1
pre-mRNA from becoming a substrate for the NMD pathway.
Cellular mRNA levels are governed by competing rates of
synthesis and decay. At the same time, mRNA decay
pathways prevent the expression of defective mRNAs. The
molecular (...truncated)