Structural basis for nuclear import of splicing factors by human Transportin 3

Proc Natl Acad Sci U S A. 2014 Feb 18;111(7):2728-33. doi: 10.1073/pnas.1320755111. Epub 2014 Jan 21.

Abstract

Transportin 3 (Tnpo3, Transportin-SR2) is implicated in nuclear import of splicing factors and HIV-1 replication. Herein, we show that the majority of cellular Tnpo3 binding partners contain arginine-serine (RS) repeat domains and present crystal structures of human Tnpo3 in its free as well as GTPase Ran- and alternative splicing factor/splicing factor 2 (ASF/SF2)-bound forms. The flexible β-karyopherin fold of Tnpo3 embraces the RNA recognition motif and RS domains of the cargo. A constellation of charged residues on and around the arginine-rich helix of Tnpo3 HEAT repeat 15 engage the phosphorylated RS domain and are critical for the recognition and nuclear import of ASF/SF2. Mutations in the same region of Tnpo3 impair its interaction with the cleavage and polyadenylation specificity factor 6 (CPSF6) and its ability to support HIV-1 replication. Steric incompatibility of the RS domain and RanGTP engagement by Tnpo3 provides the mechanism for cargo release in the nucleus. Our results elucidate the structural bases for nuclear import of splicing factors and the Tnpo3-CPSF6 nexus in HIV-1 biology.

Keywords: SR protein; Transportin-SR; host factor; importin.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Active Transport, Cell Nucleus / physiology
  • Blotting, Western
  • Cell Nucleus / metabolism*
  • Chromatography, Gel
  • Chromatography, Ion Exchange
  • Crystallography, X-Ray
  • HEK293 Cells
  • HIV-1 / metabolism
  • HIV-1 / physiology*
  • Humans
  • Immunoprecipitation
  • Models, Molecular*
  • Nuclear Proteins / metabolism*
  • Oligonucleotides / genetics
  • Protein Binding
  • Protein Conformation*
  • Virus Replication / physiology
  • X-Ray Diffraction
  • beta Karyopherins / chemistry*
  • beta Karyopherins / metabolism*
  • mRNA Cleavage and Polyadenylation Factors / metabolism

Substances

  • Nuclear Proteins
  • Oligonucleotides
  • TNPO3 protein, human
  • beta Karyopherins
  • cleavage factor Im, human
  • mRNA Cleavage and Polyadenylation Factors

Associated data

  • PDB/4C0O
  • PDB/4C0P
  • PDB/4C0Q