Nuclear-Cytoplasmic Trafficking of NTF2, the Nuclear Import Receptor for the RanGTPase, Is Subjected to Regulation

Shawn C Chafe,Jacqueline B Pierce,Dev Mangroo,Imed Eddine Gallouzi

doi:10.1371/journal.pone.0042501

Abstract

NTF2 is a cytosolic protein responsible for nuclear import of Ran, a small Ras-like GTPase involved in a number of critical cellular processes, including cell cycle regulation, chromatin organization during mitosis, reformation of the nuclear envelope following mitosis, and controlling the directionality of nucleocytoplasmic transport. Herein, we provide evidence for the first time that translocation of the mammalian NTF2 from the nucleus to the cytoplasm to collect Ran in the GDP form is subjected to regulation. Treatment of mammalian cells with polysorbitan monolaurate was found to inhibit nuclear export of tRNA and proteins, which are processes dependent on RanGTP in the nucleus, but not nuclear import of proteins. Inhibition of the export processes by polysorbitan monolaurate is specific and reversible, and is caused by accumulation of Ran in the cytoplasm because of a block in translocation of NTF2 to the cytoplasm. Nuclear import of Ran and the nuclear export processes are restored in polysorbitan monolaurate treated cells overproducing NTF2. Moreover, increased phosphorylation of a phospho-tyrosine protein and several phospho-threonine proteins was observed in polysorbitan monolaurate treated cells. Collectively, these findings suggest that nucleocytoplasmic translocation of NTF2 is regulated in mammalian cells, and may involve a tyrosine and/or threonine kinase-dependent signal transduction mechanism(s).

Highlights

Eukaryotic cells compartmentalize the DNA replication and transcription apparatus in the nucleus and the translation machinery in the cytoplasm
Treatment of S. cerevisiae with rapamycin, as well as nitrogen or amino acid deprivation has been shown to cause nuclear accumulation of mature tRNAs derived from intron-containing precursors, but not those made from intronless precursors [30,31]
Recent studies have shown that the TORC1 signaling pathway of S. cerevisiae is, in part, regulating nuclear export of mature spliced tRNAs in response to availability of nitrogen sources, as inhibition of TORC1 with rapamycin resulted in inhibition of nuclear export of spliced tRNAs [30,31]

Summary

Introduction

Eukaryotic cells compartmentalize the DNA replication and transcription apparatus in the nucleus and the translation machinery in the cytoplasm This segregation requires that exchange of molecules between the two compartments takes place across the double lipid bilayer of the nuclear envelope in order for both processes to function optimally. The nuclear envelope is perforated with large proteinaceous assemblies known as nuclear pore complexes (NPCs). These macromolecular complexes range in size from 50 MDa in yeast to 125 MDa in vertebrates [1]. Some proteins and RNAs that are smaller than the 60 kDa exclusion limit are not free to diffuse across the pore even though they are below the size restriction of the inner core; these molecules and those that are much larger in size require a carrier-mediated active transport process in order to move through the NPC

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