A conserved biogenesis pathway for nucleoporins: Proteolytic processing of a 186-kilodalton precursor generates Nup98 and the novel nucleoporin, Nup96

Beatriz M.A. Fontoura, Günter Blobel, Michael J. Matunis

Research output: Contribution to journalArticlepeer-review

Abstract

The mammalian nuclear pore complex (NPC) is comprised of ~50 unique proteins, collectively known as nucleoporins. Through fractionation of rat liver nuclei, we have isolated >30 potentially novel nucleoporins and have begun a systematic characterization of these proteins. Here, we present the characterization of Nup96, a novel nucleoporin with a predicted molecular mass of 96 kD. Nup96 is generated through an unusual biogenesis pathway that involves synthesis of a 186-kD precursor protein. Proteolytic cleavage of the precursor yields two nucleoporins: Nup98, a previously characterized GLFG- repeat containing nucleoporin, and Nup96. Mutational and functional analyses demonstrate that both the Nup98-Nup96 precursor and the previously characterized Nup98 (synthesized independently from an alternatively spliced mRNA) are proteolytically cleaved in vivo. This biogenesis pathway for Nup98 and Nup96 is evolutionarily conserved, as the putative Saccharomyces cerevisiae homologues, N-Nup145p and C-Nup145p, are also produced through proteolytic cleavage of a precursor protein. Using immunoelectron microscopy, Nup96 was localized to the nucleoplasmic side of the NPC, at or near the nucleoplasmic basket. The correct targeting of both Nup96 and Nup98 to the nucleoplasmic side of the NPC was found to be dependent on proteolytic cleavage, suggesting that the cleavage process may regulate NPC assembly. Finally, by biochemical fractionation, a complex containing Nup96, Nup107, and at least two Sec13-related proteins was identified, revealing that a major sub-complex of the NPC is conserved between yeast and mammals.

Original languageEnglish (US)
Pages (from-to)1097-1112
Number of pages16
JournalJournal of Cell Biology
Volume144
Issue number6
DOIs
StatePublished - Mar 22 1999

Keywords

  • Nuclear pore complex
  • Nucleocytoplasmic transport
  • Nucleoporin
  • Proteolytic cleavage
  • RNA export

ASJC Scopus subject areas

  • Cell Biology

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