G ₂ /M Cell Cycle Arrest Correlates with Primate Lentiviral Vpr Interaction with the SLX4 Complex

Abstract

V iral protein R (Vpr) is an accessory protein common to all primate lentiviruses whose role in viral replication remains unclear. It is packaged specifically into virions and is associated with the reverse-transcription complex during early steps of infection (1, 2). Although implicated in various virological processes, the major phenotype ascribed to Vpr is the induction of G 2 /M cell cycle arrest in dividing cells (3, 4). The importance of this phenotype in human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) replication has been notoriously difficult to define because of the minor effects of its deletion on viral replication in cell culture (5). However, reversion of inactivating mutations in both primates and humans highlights that Vpr plays an essential role for viral replication in vivo (6). In addition to cell cycle arrest, several SIV Vpr alleles are capable of targeting the host restriction factor SAMHD1 and that function became separated in the sooty mangabey SIV (SIVsm) lineage upon duplication to generate the vpx gene peculiar to this group (7). Importantly, at present HIV-1 Vpr is not known to retain any SAMHD1 counteractivity (7-10).Vpr-mediated cell cycle arrest depends on its localization to nuclear compartments and the ability to hijack a Cullin-4/DDB1/ Roc1 E3 ubiquitin ligase complex through the WD-containing DCAF-1 adaptor (11,12). For HIV-1 Vpr, this leads ultimately to the activation of the ataxia telangiectasia M and Rad3-related (ATR) and Chk1 kinases to prevent cell cycle progression into mitosis (13). Although several putative Vpr targets have been described, the identity of that which triggers this event has until recently remained elusive. The requirement for the ubiquitin-proteasome system and, in particular, K48-ubiquitin linkages supported a notion that the Vpr target is degraded (14). However, Laguette and colleagues recently found that Vpr interacts with the SLX4 complex, members of the Fanconi anemia DNA repair pathway (15). SLX4, also known as Fanconi anemia complementation group P (FANCP), is a large adaptor protein that acts as a scaffold for a heterodimeric structure-specific endonuclease comprised of MUS81 and EME1. This interaction directs this endonuclease and

SLX4-SLX1 Protein-independent Down-regulation of MUS81-EME1 Protein by HIV-1 Viral Protein R (Vpr)

“…7A). Spatial arrangement of SLX4com allows poly-ubiquitination of MUS81, mediated by CRL4-DCAF1-Vpr, resulting in its proteasome-dependent degradation (54,55). In this report, we provide evidence that MUS81-EME1 interacts with the CRL4-DCAF1 complex irrespective of Vpr and that the level of the endonuclease is regulated by Vpr binding to CRL4-DCAF1 and MUS81-EME1 (Fig.…”

“…These interactions were proposed to lead to aberrant DNA cleavage and cell cycle arrest (54). Additionally, MUS81 was reported to be down-regulated by Vpr, which was further confirmed by another two groups (55,56). However, the detailed mechanisms of MUS81 down-regulation and its relation to Vpr-induced G 2 arrest are not clear.…”

Sensing of HIV-1 Infection in Tzm-bl Cells with Reconstituted Expression of STING

“…The recent landmark study by Laguette and colleagues suggested a first molecular explanation for this activity by unraveling the interaction of Vpr with SLX4, a nuclease scaffold with important roles in DNA damage and repair (44,73). Via this interaction, Vpr prematurely activates SLX4 complexes, which results in cell cycle arrest but also reduces the proinflammatory cytokine response to HIV infection (44,59,74). As speculated by these authors, the effect of Vpr on cytokine induction may reflect a reduction of viral genomes available for sensing that results from triggered endonuclease activity of the SLX4 complex.…”

Vpr Enhances Tumor Necrosis Factor Production by HIV-1-Infected T Cells

“…The R62P mutant is unable to form nuclear foci (55). We also used mutants unable to bind proteins known to interact with Vpr, such as UNG2 (W54R) (30,105), TAK1 (S79A) (71), the nuclear envelope protein hCG1 (K27M) (30), and members of the SLX4com (Q65R, R80A) (62,63). We introduced the corresponding mutations into the HIV-1 NL4-3 provirus and measured infection and TNF levels as previously described.…”

“…Vpr arrests the cell cycle in the G 2 phase by hijacking the DCAF1-DDB1-Cul4A ubiquitin-ligase complex (56)(57)(58)(59)(60)(61). It has also been reported that the premature activation of the structure-specific endonuclease regulator SLX4 complex (SLX4com) by Vpr, through its interaction with DCAF1, mediates G 2 cell cycle arrest (62,63). The SLX4com is involved in the Fanconi anemia DNA repair pathway, thus linking the DDR with the effect of Vpr on the cell cycle.…”