RT Journal Article
SR Electronic
T1 Binding stoichiometry and structural model of the HIV-1 Rev/Importin β complex
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.11.16.468785
DO 10.1101/2021.11.16.468785
A1 Didier Spittler
A1 Rose-Laure Indorato
A1 Elisabetta Boeri Erba
A1 Elise Delaforge
A1 Luca Signor
A1 Simon J. Harris
A1 Isabel Garcia-Saez
A1 Andrés Palencia
A1 Frank Gabel
A1 Martin Blackledge
A1 Marjolaine Noirclerc-Savoye
A1 Carlo Petosa
YR 2021
UL http://biorxiv.org/content/early/2021/11/20/2021.11.16.468785.abstract
AB HIV-1 Rev mediates the nuclear export of intron-containing viral RNA transcripts and is essential for viral replication. Rev is imported into the nucleus by the host protein Importin β (Impβ), but how Rev associates with Impβ is poorly understood. Here we report biochemical, biophysical and structural studies of the Impβ/Rev complex. Gel shift, native mass spectrometry and isothermal titration calorimetry data reveal that Impβ binds two Rev monomers through independent binding sites. Small-angle X-ray scattering (SAXS) data suggest that the HEAT repeats of Impβ retain an extended conformation upon binding Rev, which according to NMR data is primarily recognized through its helical hairpin domain. Peptide scanning data and charge-reversal mutations identify the N-terminal tip of Rev helix α2 within Rev’s Arginine-Rich Motif (ARM) as a primary Impβ binding epitope. Crosslinking mass spectrometry and compensatory mutagenesis data combined with molecular docking simulations suggest a structural model in which one Rev monomer binds to the C-terminal half of Impβ with Rev helix α2 roughly parallel to the HEAT-repeat superhelical axis while the other monomer binds to the N-terminal half. These findings shed light on the molecular basis of Rev recognition by Impβ and highlight an atypical binding behaviour that distinguishes Rev from canonical cellular Impβ cargos.Competing Interest StatementThe authors have declared no competing interest.