Multiple Conserved Domains of the Nucleoporin Nup124p and Its Orthologs Nup1p and Nup153 Are Critical for Nuclear Import and Activity of the Fission Yeast Tf1 Retrotransposon

Abstract

The nucleoporin Nup124p is a host protein required for the nuclear import of both, retrotransposon Tf1-Gag as well as the retroviral HIV-1 Vpr in fission yeast. The human nucleoporin Nup153 and the Saccharomyces cerevisiae Nup1p were identified as orthologs of Nup124p. In this study, we show that all three nucleoporins share a large FG/FXFG-repeat domain and a C-terminal peptide sequence, GRKIxxxxxRRKx, that are absolutely essential for Tf1 retrotransposition. Though the FXFG domain was essential, the FXFG repeats themselves could be eliminated without loss of retrotransposon activity, suggesting the existence of a common element unrelated to FG/FXFG motifs. The Nup124p C-terminal peptide, GRKIAVPRSRRKR, was extremely sensitive to certain single amino acid changes within stretches of the basic residues. On the basis of our comparative study of Nup124p, Nup1p, and Nup153 domains, we have developed peptides that specifically knockdown retrotransposon activity by disengaging the Tf1-Gag from its host nuclear transport machinery without any harmful consequence to the host itself. Our results imply that those domains challenged a specific pathway affecting Tf1 transposition. Although full-length Nup1p or Nup153 does not complement Nup124p, the functionality of their conserved domains with reference to Tf1 activity suggests that these three proteins evolved from a common ancestor.

The Nup153-Nup50 Protein Interface and Its Role in Nuclear Import

“…In Nup124p (Schizosaccharomyces pombe), this C-terminal motif was found to be critical retrotransposon activity, specifically for Tf1-Gag import (29). Interestingly, the other region of Nup124p critical to retrotransposon function shares homology with Nup153 as well, in this case, at the site within the N-terminal domain of Nup153 (residues 448 -634) that we found here is important for the interface with Nup50 (35).…”

“…The C-terminal importin ␣-docking site on Nup153 is highly conserved across species (29) and has affinity for a range of importin ␣ isoforms (supplemental Fig. S3) (29).…”

“…S3) (29). In the context of the Saccharomyces cerevisiae basket nucleoporin Nup1p, this tail region has been found to be important for the efficiency of import (30).…”

Introgression of Drosophila simulans Nuclear Pore Protein 160 in Drosophila melanogaster Alone Does Not Cause Inviability but Does Cause Female Sterility

“…Why do NPC protein genes evolve so quickly at the molecular level? The nuclear entry of retroviruses and retrotransposons is mediated by NPC (e.g., Dang and Levin 2000;Petersen et al 2000;von Kobbe et al 2000;Gustin and Sarnow 2001;Sistla et al 2007;Beliakova-Bethell et al 2009), and this host-pathogen interaction may have accelerated the evolution of NPC proteins. Alternatively, the NPC proteins may have evolved so quickly to interact properly with rapidly evolving sequences of the centromere and its surrounding heterochromatin (Sawamura et al 1993b;Henikoff et al 2001;Bayes and Malik 2009;Ferree and Barbash 2009).…”

Flexible Gates: Dynamic Topologies and Functions for FG Nucleoporins in Nucleocytoplasmic Transport

“…Even within a single FG domain there is evidence for redundancy among FG repeats. For example, the Tf1 retrotransposon of fission yeast requires FXFG repeats within Nup124 for nuclear import but shows no discernible preference for any single repeat within the FXFG domain (151).…”