Bishop lab

Retroviral Replication Laboratory

: The function of the HIV accessory proteins, Vpx and Vpr

Electron microscope image of mouse leukaemia virus-like particles containing a faulty p12 protein.

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The HIV accessory proteins Vpx and Vpr are not required for replication in many cell lines but are important in vivo and, as such, have been evolutionary conserved. We are investigating how these proteins enhance HIV infection.

Vpx and Vpr proteins

Figure 1: A), Dimeric inactive SAMHD1 (blue) binds dGTP at the dimer interface. Activated SAMHD1 (red) catalyses the cleavage of dNTPs into the composite deoxynucleoside and inorganic triphosphate. SAMHD1 activity in myeloid cells suppresses the deoxynucleotide pool, inhibiting reverse transcriptase and blocking infection by HIV-1. (B) In the presence of Vpx, SAMHD1 is recruited by the DDB-CUL4-DCAF1 E3 ubiquitin ligase complex and targeted to the proteasome for degradation. Control on the dNTP pool is released and sufficient dNTPs are available for reverse transcription to be completed, allowing infection by HIV-2 and SIVs that encode Vpx. (C) AGS mutations in the allosteric binding site of SAMHD1 prevent dGTP binding or allosteric activation rendering the protein inactive. As a result, deoxynucleotide levels rise and aberrant DNA products arising from reverse transcription of endogenous retroviruses accumulate within the cytoplasm, triggering the inappropriate production of interferon observed in AGS.

The HIV-2 protein Vpx enhances replication of both HIV-1 and HIV-2 in cells of the myeloid lineage and resting T cells. In 2011, the target for Vpx was identified as the cellular protein SAMHD1 (sterile alpha motif and an histidine-aspartate domain-containing protein 1). Vpx recruits SAMHD1 to the DDB1/CUL4A/ROC1 E3 ubiquitin ligase complex through interaction with the substrate-adaptor protein DCAF1 resulting in proteasomal degradation of SAMHD1. Mutations in SAMHD1 in humans lead to Aicardi Goutières syndrome (AGS), which mimics congenital viral infection. Ian Taylor's group at the Crick demonstrated that SAMHD1 is a GTP/dGTP-activated deoxynucleotide triphosphohydrolase that degrades dNTPs to constituent nucleoside and inorganic triphosphate. This led to the hypothesis that SAMHD1 inhibited HIV-1 replication by reducing cellular dNTP pools in differentiated cells below the threshold required for reverse transcription, although alternative mechanisms have also been proposed. In collaboration with the Taylor group, we have been pursuing how SAMHD1 inhibits HIV-1 replication and investigating how Vpx recruits SAMHD1 and targets it for degradation.

Vpr is a paralogue of Vpx with an unknown function in HIV-1. Expression of Vpr induces cell-cycle arrest, and, as Vpr binds the substrate-adaptor protein DCAF1, it seems likely this is due to Vpr targeting a cellular protein for degradation. Several Vpr targets have been proposed, although none appear responsible for the cell cycle block, and there is no mechanism for how they inhibit HIV-1 replication. In addition, Vpr proteins from some SIV strains can induce degradation of SAMHD1. Continuing our work on SAMHD1 and Vpx, we are now focusing on deducing the function of Vpr. We are using cutting edge techniques to identify novel cellular target(s) for Vpr and reveal their antiviral function(s). Furthermore, we are investigating the specificity and evolutionary conservation of several recently reported Vpr interactions using our established cell-based degron assay, in vitro assays and structural studies in collaboration with Ian Taylor.