Structural and functional analysis of an essential nucleoporin heterotrimer on the cytoplasmic face of the nuclear pore complex.

So far, only a few of the interactions between the ≈ 30 nucleoporins comprising the modular structure of the nuclear pore complex have been defined at atomic resolution. Here we report the crystal structure, at 2.6 Å resolution, of a heterotrimeric complex, composed of fragments of three cytoplasmically oriented nucleoporins of yeast: Nup82, Nup116, and Nup159. Our data show that the Nup82 fragment, representing more than the N-terminal half of the molecule, folds into an extensively decorated, seven-bladed ß-propeller that forms the centerpiece of this heterotrimeric complex and anchors both a C-terminal fragment of Nup116 and the C-terminal tail of Nup159. Binding between Nup116 and Nup82 is mutually reinforced via two loops, one emanating from the Nup82 ß-propeller and the other one from the ß-sandwich fold of Nup116, each contacting binding pockets in their counterparts. The Nup82-Nup159 interaction occurs through an amphipathic α-helix of Nup159, which is cradled in a large hydrophobic groove that is generated from several large surface decorations of the Nup82 ß-propeller. Although Nup159 and Nup116 fragments bind to the Nup82 ß-propeller in close vicinity, there are no direct contacts between them, consistent with the noncooperative binding that was detected biochemically. Extensive mutagenesis delineated hot-spot residues for these interactions. We also showed that the Nup82 ß-propeller binds to other yeast Nup116 family members, Nup145N, Nup100 and to the mammalian homolog, Nup98. Notably, each of the three nucleoporins contains additional nuclear pore complex binding sites, distinct from those that were defined here in the heterotrimeric Nup82•Nup159•Nup116 complex.

Characterization of the role of the tumor marker Nup88 in mitosis.

Nuclear pore complexes are massive multiprotein channels responsible for traffic between the nucleus and cytoplasm, and are composed of approximately 30 proteins, termed nucleoporins (Nup). Our recent studies indicated that the nucleoporins Rae1 and Tpr play critical roles in maintaining the spindle bipolarity during cell division. In the present study, we found that another nucleoporin, Nup88, was localized on the spindles together with Nup214 during mitosis. Nup88 expression is linked to the progression of carcinogenesis, Nup88 has been proposed as a tumor marker. Overexpression of Nup88 enhanced multinucleated cell formation. RNAi-mediated knockdown of Nup88 disrupted Nup214 expression and localization and caused multipolar spindle phenotypes. Our data indicate that proper expression of Nup88 is critical for preventing aneuploidy formation and tumorigenesis.