The Nuclear Pore Complex and the DEAD Box Protein Rat8p/Dbp5p Have Nonessential Features Which Appear To Facilitate mRNA Export following Heat Shock

Abstract

Nuclear pore complexes (NPCs) play an essential role in RNA export. Nucleoporins required for mRNA export in Saccharomyces cerevisiae are found in the Nup84p and Nup82p subcomplexes of the NPC. The Nup82p subcomplex contains Nup82p, Rat7p/Nup159p, Nsp1p, Gle1p/Rss1p, and Rip1p/Nup42p and is found only on the cytoplasmic face of NPCs. Both Rat7p and Gle1p contain binding sites for Rat8p/Dbp5p, an essential DEAD box protein and putative RNA helicase. Rip1p interacts directly with Gle1p and is the only protein known to be essential for mRNA export after heat shock but not under normal growth conditions. We report that in cells lacking Rip1p, both Gle1p and Rat8p dissociate from NPCs following heat shock at 42°C. Rat8p but not Gle1p was retained at NPCs if rip1Δ cells were first shifted to 37°C and then to 42°C, and this was correlated with preserving mRNA export in heat-shocked rip1Δ cells. Export following ethanol shock was less dependent on the presence of Rip1p. Exposure to 10% ethanol led to dissociation of Rat8p from NPCs in both wild-type and rip1Δ cells. Following this treatment, Rat8p was primarily nuclear in wild-type cells but primarily cytoplasmic in rip1Δ cells. We also determined that efficient export of heat shock mRNA after heat shock depends upon a novel 6-amino-acid element within Rat8p. This motif is not required under normal growth conditions or following ethanol shock. These studies suggest that the molecular mechanism responsible for the defect in export of heat shock mRNAs in heat-shocked rip1Δ cells is dissociation of Rat8p from NPCs. These studies also suggest that both nuclear pores and Rat8p have features not required for mRNA export in growing cells but which enhance the ability of mRNAs to be exported following heat shock.

We thank Francoise Stutz for providing strains. We thank the Cole laboratory for helpful discussions and Catherine Heath for critically reading the manuscript.

This research was supported by Public Health Service grant GM33998 to C.N.C. from the National Institute of General Medical Sciences, National Institutes of Health.