Nuclear export of proteins in plants: AtXPO1 is the export receptor for leucine‐rich nuclear export signals in Arabidopsis thaliana

Abstract

SummaryTransport across the nuclear envelope is mediated by transport receptors from the Importin b family. We identi®ed Exportin 1 from Arabidopsis (AtXPO1/AtCRM1) as the nuclear export receptor for proteins carrying leucine-rich nuclear export signals (NESs). AtXPO1 shares 42±50% identity with its functional homologues from humans and yeasts. We functionally characterised AtXPO1 by its interaction with NESs of animal and plant proteins, which is inhibited by the cytotoxin leptomycin B (LMB), and also by its interaction with the small GTPase Ran1 in the yeast two-hybrid system. Furthermore, we demonstrated the existence of a nuclear export pathway for proteins in plants. For the characterisation of nuclear export activities, we established an in vivo assay based on the localisation equilibrium of a GFP reporter protein fused to both a nuclear localisation signal (NLS) and an NES motif. Using this in vivo assay we demonstrated that the NES of the heterologous protein Rev is also functional in plants and that its export is inhibited by LMB. In addition, we identi®ed a leucine-rich NES in the Arabidopsis protein AtRanBP1a. The NES, which is located at the carboxy terminus of the protein, is disrupted by mutating three long chain hydrophobic amino acid residues to alanine (L 176 A, L 179 A, V 181 A). In BY-2 protoplasts the NES of AtRanBP1a is functionally indistinguishable from the Rev NES. Our results demonstrate that the machinery for the nuclear export of proteins is functionally conserved in plants.

BLADE-ON-PETIOLE–Dependent Signaling Controls Leaf and Floral Patterning in Arabidopsis

“…No band corresponding to free GFP was observed in these extracts ( Figure 5B). Therefore, we concluded that BOP2-GFP is targeted to the nucleus despite lacking a canonical nuclear localization sequence, because the predicted size of BOP2-GFP (;88 kD) well exceeds the estimated size exclusion limit (;60 kD) for passive diffusion of proteins through nuclear pores (Haasen et al, 1999). A similar pattern of subcellular localization was observed for NPR1-GFP transgenic seedlings in uninduced guard cells.…”

Nuclear retention of the transcription factor NLP7 orchestrates the early response to nitrate in plants

“…In N-starved NLP7-GFP seedlings that had been treated for 3 h with leptomycin B (LMB), an inhibitor of Exportin1 (ref. 13), NLP7-GFP exhibited partial nuclear accumulation (Fig. 1i, Supplementary Fig.…”

Nucleocytoplasmic Distribution Is Required for Activation of Resistance by the Potato NB-LRR Receptor Rx1 and Is Balanced by Its Functional Domains

“…The addition of these targeting signals does not fully exclude the protein from either compartment, but enhances the active transport into (NLS) or out of the nucleus (NES) and is thereby expected to shift the balance between the nuclear and cytoplasmic pools of Rx1. To redirect Rx1 to the nucleus, the well-known SV40 Large T-antigen monopartite NLS was chosen (Lanford and Butel, 1984;Haasen et al, 1999). The PKI nuclear export signal (Wen et al, 1995) was applied to direct Rx1 from the nucleus to the cytoplasm.…”

“…The classical monopartite or bipartite nuclear localization signals (NLSs) are well defined and consist of short stretches of basic residues (three to six Lys/Arg) that interact with specific binding surfaces on the import receptor importin a, which in turn forms a heterodimer with importin b (Gö rlich and Kutay, 1999). Nuclear export is directed via Leu-rich nuclear export signals (NESs), which interact with the nuclear export receptor (exportin) (Haasen et al, 1999;Hutten and Kehlenbach, 2007). Several NB-LRR proteins, like RRS1-R, RPS4, and SNC1, do contain functional NLS sequences (Deslandes et al, 2003;Wirthmueller et al, 2007;Cheng et al, 2009).…”