A nuclear role for the DEAD-box protein Dbp5 in tRNA export.

Azra Lari,Barry P Young,Rachel Montpetit,Ben Montpetit,Taylor Reiter,Rima Sandhu,Arvind Arul Nambi Rajan,Chris JR Loewen

doi:10.7554/elife.48410

Azra Lari, Barry P Young + Show 6 more

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https://doi.org/10.7554/elife.48410

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Abstract

Dbp5 is an essential DEAD-box protein that mediates nuclear mRNP export. Dbp5 also shuttles between nuclear and cytoplasmic compartments with reported roles in transcription, ribosomal subunit export, and translation; however, the mechanism(s) by which nucleocytoplasmic transport occurs and how Dbp5 specifically contributes to each of these processes remains unclear. Towards understanding the functions and transport of Dbp5 in Saccharomyces cerevisiae, alanine scanning mutagenesis was used to generate point mutants at all possible residues within a GFP-Dbp5 reporter. Characterization of the 456 viable mutants led to the identification of an N-terminal Xpo1-dependent nuclear export signal in Dbp5, in addition to other separation-of-function alleles, which together provide evidence that Dbp5 nuclear shuttling is not essential for mRNP export. Rather, disruptions in Dbp5 nucleocytoplasmic transport result in tRNA export defects, including changes in tRNA shuttling dynamics during recovery from nutrient stress.

Highlights

In eukaryotic cells, spatial separation of the transcriptional and translational processes necessitates transport of RNA and protein cargos across the nuclear envelope (NE) via nuclear pore complexes (NPCs)
25 alanine substitutions were identified as lethal (Supplementary file 1 - Table 1)
Lethal mutations mapped to regions of Dbp5 that participate in binding NPC regulators Gle1 and Nup159, including residues R256, Y325, and K382 (Dossani et al, 2009; Montpetit et al, 2011; Weirich et al, 2004)

Summary

Introduction

Spatial separation of the transcriptional and translational processes necessitates transport of RNA and protein cargos across the nuclear envelope (NE) via nuclear pore complexes (NPCs). Non-coding RNA (ncRNA) species, such as ribosomal RNA (rRNA) or transfer RNA (tRNA), undergo nuclear processing and export following transcription (Hopper, 2013; Zemp and Kutay, 2007). Throughout these events, RNA-binding proteins (RBPs) interact with each RNA to form a ribonucleoprotein complex (RNP), with the protein constituents of each RNP mediating RNA processing and RNP functions within the cell (Moore, 2005).

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