The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA

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Abstract

The HIV Rev protein routes viral RNAs containing the Rev Response Element (RRE) through the Crm1 nuclear export pathway to the cytoplasm where viral proteins are expressed and genomic RNA is delivered to assembling virions. The RRE assembles a Rev oligomer that displays nuclear export sequences (NESs) for recognition by the Crm1-Ran^GTP nuclear receptor complex. Here we provide the first view of an assembled HIV-host nuclear export complex using single-particle electron microscopy. Unexpectedly, Crm1 forms a dimer with an extensive interface that enhances association with Rev-RRE and poises NES binding sites to interact with a Rev oligomer. The interface between Crm1 monomers explains differences between Crm1 orthologs that alter nuclear export and determine cellular tropism for viral replication. The arrangement of the export complex identifies a novel binding surface to possibly target an HIV inhibitor and may point to a broader role for Crm1 dimerization in regulating host gene expression.

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David S Booth

Graduate Group in Biophysics, University of California, San Francisco, San Francisco, United States

Competing interests
The authors declare that no competing interests exist.
Yifan Cheng

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

Competing interests
The authors declare that no competing interests exist.
Alan D Frankel

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

For correspondence
frankel@cgl.ucsf.edu

Competing interests
The authors declare that no competing interests exist.

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This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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David S Booth
Yifan Cheng
Alan D Frankel

(2014)

The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA

eLife 3:e04121.

https://doi.org/10.7554/eLife.04121

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Human
Virus

1. Biochemistry and Chemical Biology
2. Microbiology and Infectious Disease
RNA-directed remodeling of the HIV-1 protein Rev orchestrates assembly of the Rev–Rev response element complex

Bhargavi Jayaraman, David C Crosby ... Alan D Frankel

Research Article Updated Jan 9, 2015
The HIV-1 protein Rev controls a critical step in viral replication by mediating the nuclear export of unspliced and singly-spliced viral mRNAs. Multiple Rev subunits assemble on the Rev Response Element (RRE), a structured region present in these RNAs, and direct their export through the Crm1 pathway. Rev-RRE assembly occurs via several Rev oligomerization and RNA-binding steps, but how these steps are coordinated to form an export–competent complex is unclear. Here, we report the first crystal structure of a Rev dimer-RRE complex, revealing a dramatic rearrangement of the Rev-dimer upon RRE binding through re-packing of its hydrophobic protein–protein interface. Rev-RNA recognition relies on sequence-specific contacts at the well-characterized IIB site and local RNA architecture at the second site. The structure supports a model in which the RRE utilizes the inherent plasticity of Rev subunit interfaces to guide the formation of a functional complex.
1. Microbiology and Infectious Disease
Rev Protein: Really exasperating viral protein from HIV

James R Williamson

Insight Jan 9, 2015
Two new structures shed additional light on the nuclear transport of viral transcripts.
1. Structural Biology and Molecular Biophysics
Water and chloride as allosteric inhibitors in WNK kinase osmosensing

Liliana R Teixeira, Radha Akella ... Elizabeth J Goldsmith

Research Article Nov 25, 2024
Osmotic stress and chloride regulate the autophosphorylation and activity of the WNK1 and WNK3 kinase domains. The kinase domain of unphosphorylated WNK1 (uWNK1) is an asymmetric dimer possessing water molecules conserved in multiple uWNK1 crystal structures. Conserved waters are present in two networks, referred to here as conserved water networks 1 and 2 (CWN1 and CWN2). Here, we show that PEG400 applied to crystals of dimeric uWNK1 induces de-dimerization. Both the WNK1 the water networks and the chloride-binding site are disrupted by PEG400. CWN1 is surrounded by a cluster of pan-WNK-conserved charged residues. Here, we mutagenized these charges in WNK3, a highly active WNK isoform kinase domain, and WNK1, the isoform best studied crystallographically. Mutation of E314 in the Activation Loop of WNK3 (WNK3/E314Q and WNK3/E314A, and the homologous WNK1/E388A) enhanced the rate of autophosphorylation, and reduced chloride sensitivity. Other WNK3 mutants reduced the rate of autophosphorylation activity coupled with greater chloride sensitivity than wild-type. The water and chloride regulation thus appear linked. The lower activity of some mutants may reflect effects on catalysis. Crystallography showed that activating mutants introduced conformational changes in similar parts of the structure to those induced by PEG400. WNK activating mutations and crystallography support a role for CWN1 in WNK inhibition consistent with water functioning as an allosteric ligand.