ABSTRACT
Nuclear pore complexes (NPCs) facilitate macromolecular exchange between the nucleus and cytoplasm of eukaryotic cells. The vertebrate NPC is composed of approximately 30 different proteins (nucleoporins), of which around one third contain phenylalanine-glycine (FG)-repeat domains that are thought to mediate the main interaction between the NPC and soluble transport receptors. We have recently shown that the FG-repeat domain of Nup153 is flexible within the NPC, although this nucleoporin is anchored to the nuclear side of the NPC. By using domain-specific antibodies, we have now mapped the domain topology of Nup214 in Xenopus oocytes and in human somatic cells by immuno-EM. We have found that whereas Nup214 is anchored to the cytoplasmic side of the NPC via its N-terminal and central domain, its FG-repeat domain appears flexible, residing on both sides of the NPC. Moreover, the spatial distribution of the FG-repeat domains of both Nup153 and Nup214 shifts in a transport-dependent manner, suggesting that the location of FG-repeat domains within the NPC correlates with cargo/receptor interactions and that they concomitantly move with cargo through the central pore of the NPC.
Subject(s)
Nuclear Pore Complex Proteins/chemistry , Nuclear Pore/metabolism , Animals , Antibody Specificity , Biological Transport, Active , Female , HL-60 Cells , HeLa Cells , Humans , In Vitro Techniques , Microscopy, Immunoelectron , Models, Molecular , Nuclear Pore/ultrastructure , Nuclear Pore Complex Proteins/genetics , Nuclear Pore Complex Proteins/immunology , Nuclear Pore Complex Proteins/metabolism , Oocytes/metabolism , Protein Structure, Tertiary , Rabbits , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Recombinant Proteins/metabolism , Repetitive Sequences, Amino Acid , Xenopus laevisABSTRACT
Most nucleocytoplasmic traffic through the nuclear pore complex is mediated by soluble receptors of the importin/exportin or karyopherin family. mRNA export is unique in that no receptor of this family has been implicated in trafficking of the bulk of mRNAs. Instead, many diverse proteins have been linked to mRNA export, but an all-encompassing model remains elusive. Understanding how these proteins interact with each other is central to the development of such a model. Here, we have focused on the interactions between three proteins implicated in mRNA export, Nup98, Rae1/Gle2, and TAP. We have defined the binary complexes that form among these proteins. We find that Gle2 requires two sites within TAP for stable interaction. Strikingly, rather than a general affinity for all nucleoporin FG repeats, TAP has highest affinity for a specific region within the GLFG domain of Nup98, indicating that not all repeats are identical in function. We have established that the ternary complex can form through simultaneous binding of both Gle2 and TAP to adjacent sites on Nup98. In contrast, Nup98 competes with TAP for Gle2 binding; when bound to Nup98, Gle2 no longer interacts directly with TAP. From these interactions, we propose that Gle2 may act to deliver TAP to Nup98 and that this may represent the first in a series of interactions between an export complex and a nucleoporin.
Subject(s)
ATP-Binding Cassette Transporters/metabolism , Nuclear Matrix-Associated Proteins/metabolism , Nuclear Pore Complex Proteins/metabolism , Nucleocytoplasmic Transport Proteins/metabolism , RNA, Messenger/metabolism , ATP-Binding Cassette Transporters/chemistry , Active Transport, Cell Nucleus/physiology , Animals , Binding Sites , Cloning, Molecular , Nuclear Matrix-Associated Proteins/genetics , Nuclear Pore/metabolism , Nucleocytoplasmic Transport Proteins/genetics , XenopusABSTRACT
Almost 50 years after its discovery, a cranium from the early Miocene of Rusinga Island, Kenya, is designated the type specimen for a new lorisid species in the genusMioeuoticus. This new species differs fromMioeuoticus bishopi in a number of dental attributes. Several cranial features place this species in Lorisidae, where it may represent the sister group to living lorises.
