NUP93, A VERTEBRATE HOMOLOG OF YEAST NIC96P, FORMS A COMPLEX WITH A NOVEL 205-KDA PROTEIN AND IS REQUIRED FOR CORRECT NUCLEAR-PORE ASSEMBLY

Yeast and vertebrate nuclear pores display significant morphological s imilarity by electron microscopy, but sequence similarity between the respective proteins has been more difficult to observe. Herein we have identified a vertebrate nucleoporin, Nup93, in both human and Xenopus that has proved to be an evolutionarily related homologue of the yeas t nucleoporin Nic96p. Polyclonal antiserum to human Nup93 detects corr esponding proteins in human rat, and Xenopus cells. Immunofluorescence and immunoelectron microscopy localize vertebrate Nup93 at the nuclea r basket and at or near the nuclear entry to the gated channel of the pore. Immunoprecipitation from both mammalian and Xenopus cell extract s indicates that a small fraction of Nup93 physically interacts with t he nucleoporin p62, just as yeast Nic96p interacts with the yeast p62 homologue. However, a large fraction of vertebrate Nup93 is extracted from pores and is also present in Xenopus egg extracts in complex with a newly discovered 205-kDa protein. Mass spectrometric sequencing of the human 205-kDa protein reveals that this protein is encoded by an o pen reading frame, KIAO225, present in the human database. The putativ e human nucleoporin of 205kDa has related sequence homologues in Caeno rhabditis elegans and Saccharomyces cerevisiae. To analyze the role of the Nup93 complex in the pore, nuclei were assembled that lack the Nu p93 complex after immunodepletion of a Xenopus nuclear reconstitution extract. The Nup93-complex-depleted nuclei are clearly defective for c orrect nuclear pore assembly. From these experiments, we conclude that the vertebrate and yeast pore have significant homology in their func tionally important cores and that, with the identification of Nup93 an d the 205-kDa protein, we have extended the knowledge of the nearest-n eighbor interactions of this core in both yeast and vertebrates.