- © 2010 American Society of Plant Biologists
Getting molecules in and out of the nucleus is a critical function of eukaryotic cells. The task is performed by the nuclear pore complex (NPC), a massive protein complex (estimated at 125 MD in vertebrates) that spans the nuclear envelope and allows diffusion of small molecules but tightly controls transport of larger molecules, such as proteins, RNA, and ribosomal subunits. The NPC comprises at least 30 different proteins, called nucleoporins. Knowledge of the functions of individual components and the overall structure in plants lags behind that of vertebrates and yeast, for which the NPC is well characterized (recently reviewed in Wälde and Kehlenbach, 2010). Tamura et al. (pages 4084–4097) present a detailed investigation of components of the NPC in Arabidopsis based on an interactive proteomic approach using green fluorescent protein (GFP)-tagged nucleoporins.
The authors started with a known plant nucleoporin, RNA export factor 1 (RAE1), which is encoded by numerous plant genomes. Transgenic Arabidopsis plants expressing GFP-tagged At-RAE1 were generated and used to analyze the NPC in living cells and to identify other nucleoporins that make up the NPC in Arabidopsis. Nucleoporins were identified by immunoprecipation from transgenic Arabidopsis tissue using anti-GFP antibody followed by mass spectrometry identification of proteins present in the immunoprecipitate. Using this approach, eight known plant nucleoporins and 22 novel nucleoporins were isolated and further characterized by cloning and localization of GFP-nucleoporin fusion proteins.
Most of the Arabidopsis nucleoporins show homology to human nucleoporins, except for one protein, called Nup136/Nup1, which was found to be plant specific. Database searches revealed Nup136/Nup1 homologs in a number of plant species but not in yeast or vertebrates. T-DNA insertional mutants of nup136/nup1 showed defects in flowering time and pollen development (see figure), suggesting that Nup136 plays a role in aspects of flowering and reproductive development in plants. Structural features of Nup136/Nup1 indicate that the protein might be involved in regulating active transport through the NPC.
Mutants of nup136 exhibit defects in plant development. Top panel: nup136 knockout mutants showed a defect in fruit maturation. Inset shows a magnified view of representative siliques. Bottom panel: Scanning electron micrographs of pollen grains of wild-type (wt) and nup136 mutant plants. Arrows indicate the shrunken pollen grains of the mutants. Bar = 50 μm. (Figure reproduced from Tamura et al. [2010].)
Overall, the Arabidopsis nucleoporins identified shared higher homology with vertebrate than with yeast nucleoporins, suggesting that the plant NPC structure is more similar to that of vertebrates than yeast. This work shows that the basic NPC structure is conserved in plants, yeast, and vertebrates, although each organism uses specialized nucleoporins with functions specific to their own NPCs. The identification of 22 novel plant nucleoporins will facilitate further studies into their interactions, regulatory roles, and biological functions within the NPC.
