Abstract
Nuclear mRNA export is a crucial step in eukaryotic gene expression, which is in yeast coupled to cotranscriptional messenger ribonucleoprotein particle (mRNP) assembly and surveillance. Several surveillance systems that monitor nuclear mRNP biogenesis and export have been described, but the mechanism by which the improper mRNPs are recognized and eliminated remains poorly understood. Here we report that the conserved PIN domain protein Swt1 is an RNA endonuclease that participates in quality control of nuclear mRNPs and can associate with the nuclear pore complex (NPC). Swt1 showed endoribonuclease activity in vitro that was inhibited by a point mutation in the predicted catalytic site. Swt1 lacked clear sequence specificity but showed a strong preference for single-stranded regions. Genetic interactions were found between Swt1 and the THO/TREX and TREX-2 complexes, and with components of the perinuclear mRNP surveillance system, Mlp1, Nup60, and Esc1. Inhibition of the nuclease activity of Swt1 increased the levels and cytoplasmic leakage of unspliced aberrant pre-mRNA, and induced robust nuclear poly(A)+ RNA accumulation in mlp1Δ and esc1Δ strains. Overexpression of Swt1 also caused strong nuclear poly(A)+ RNA accumulation. Swt1 is normally distributed throughout the nucleus and cytoplasm but becomes concentrated at nuclear pore complexes (NPCs) in the nup133Δ mutant, which causes NPC clustering and defects in mRNP export. The data suggest that Swt1 endoribonuclease might be transiently recruited to NPCs to initiate the degradation of defective pre-mRNPs or mRNPs trapped at nuclear periphery in order to avoid their cytoplasmic export and translation.
Highlights
Nuclear export of messenger RNA is a fundamental process of eukaryotic gene expression
To avoid nuclear export of aberrant transcripts and their translation in the cytoplasm, the quality of nuclear messenger ribonucleoprotein particle (mRNP) is monitored by several surveillance systems
We show that the conserved protein Swt1 is an RNA endoribonuclease, an RNA-degrading enzyme, that becomes indispensable when factors involved in co-transcriptional mRNP assembly and mRNP quality control are mutated
Summary
Nuclear export of messenger RNA (mRNA) is a fundamental process of eukaryotic gene expression. Newly synthesized pre-mRNAs are cotranscriptionally assembled into messenger ribonucleoprotein particles (mRNPs) with the assistance of transcription-export (TREX) complexes. In the absence of TREX or TREX-2 components, nascent mRNAs are not correctly assembled into mRNPs, which can lead to hybridization with the DNA template (R-loops formation), defective polyadenylation, and accumulation at the 39 end of the gene [3,4,5]. Nuclear mRNPs are monitored by surveillance systems to prevent the export and subsequent translation of aberrant mRNAs (reviewed in [8,9]). In TREX mutants, imperfect mRNPs can be targeted and degraded by the nuclear exosome together with the TRAMP polyadenylation complex [10]
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