Structure of HIV-1 Vpr in complex with the human nucleotide excision repair protein hHR23A

In-Ja L Byeon,Maria Delucia,Chang H Byeon,Jinwoo Ahn,Ying Wu,Guillermo Calero,Caili Hao,Jinwon Jung,Jacek Skowronski,Simon Weiss,Angela M Gronenborn,Xiaohong Zhou

doi:10.1038/s41467-021-27009-w

Abstract

HIV-1 Vpr is a prototypic member of a large family of structurally related lentiviral virulence factors that antagonize various aspects of innate antiviral immunity. It subverts host cell DNA repair and protein degradation machineries by binding and inhibiting specific post-replication repair enzymes, linking them via the DCAF1 substrate adaptor to the Cullin 4 RING E3 ligase (CRL4DCAF1). HIV-1 Vpr also binds to the multi-domain protein hHR23A, which interacts with the nucleotide excision repair protein XPC and shuttles ubiquitinated proteins to the proteasome. Here, we report the atomic resolution structure of Vpr in complex with the C-terminal half of hHR23A, containing the XPC-binding (XPCB) and ubiquitin-associated (UBA2) domains. The XPCB and UBA2 domains bind to different sides of Vpr’s 3-helix-bundle structure, with UBA2 interacting with the α2 and α3 helices of Vpr, while the XPCB domain contacts the opposite side of Vpr’s α3 helix. The structure as well as biochemical results reveal that hHR23A and DCAF1 use overlapping binding surfaces on Vpr, even though the two proteins exhibit entirely different three-dimensional structures. Our findings show that Vpr independently targets hHR23A- and DCAF1- dependent pathways and highlight HIV-1 Vpr as a versatile module that interferes with DNA repair and protein degradation pathways.

Highlights

HIV-1 Vpr is a prototypic member of a large family of structurally related lentiviral virulence factors that antagonize various aspects of innate antiviral immunity
Repair and chromatin metabolism, including the base excision repair (BER) protein Uracil-DNA glycosylase (UNG2)[14,15], the double strand break repair and Holliday junction resolvase MUS81-EME116,17, the helicase-like transcription factor (HLTF) that plays a role in the repair of stalled replication forks[18,19,20], Exo[1], a nuclease involved in double strand break repair[21], and
In order to prepare a homogeneous sample of the complex, we generated Vpr[1–79] and hHR23A223–363 fusion constructs, connecting Vpr to either the N- or C-terminus of the double domain construct of hHR23A by a six amino acid (GGS)[2] linker (L)

Summary

Introduction

HIV-1 Vpr is a prototypic member of a large family of structurally related lentiviral virulence factors that antagonize various aspects of innate antiviral immunity It subverts host cell DNA repair and protein degradation machineries by binding and inhibiting specific post-replication repair enzymes, linking them via the DCAF1 substrate adaptor to the Cullin 4 RING E3 ligase (CRL4DCAF1). Vpr binds to DCAF1 and mediates loading of different protein substrates onto this E3 ligase, leading to their polyubiquitination and proteasome-dependent degradation[11,12,13] Among these substrates are proteins involved in post-replication DNA repair and chromatin metabolism, including the base excision repair (BER) protein Uracil-DNA glycosylase (UNG2)[14,15], the double strand break repair and Holliday junction resolvase MUS81-EME116,17, the helicase-like transcription factor (HLTF) that plays a role in the repair of stalled replication forks[18,19,20], Exo[1], a nuclease involved in double strand break repair[21], and TET2, a methylcytosine dioxygenase that is a potent epigenetic regulator of gene expression in hematopoietic cells[22]. In contrast to other DNA repair proteins that are targeted by Vpr, hHR23A does not become ubiquitinated in a Vpr-dependent manner and is not depleted in cells expressing HIV-1 Vpr[21], suggesting that hHR23A is not a Vpr-recruited CRL4DCAF1 E3 substrate and that the Vpr–hHR23A complex likely presents a distinct architecture

Methods

Results

Conclusion