Aicardi-Goutieres syndrome gene and HIV-1 restriction factor SAMHD1 is a dGTP-regulated deoxynucleotide triphosphohydrolase

Abstract

The SAMHD1 protein is an HIV-1 restriction factor that is targeted by the HIV-2 accessory protein Vpx in myeloid lineage cells. Mutations in the SAMHD1 gene cause Aicardi-Goutières syndrome, a genetic disease that mimics congenital viral infection. To determine the physiological function of the SAMHD1 protein, the SAMHD1 gene was cloned, recombinant protein was produced, and the catalytic activity of the purified enzyme was identified. We show that SAMHD1 contains a dGTP-regulated deoxynucleotide triphosphohydrolase. We propose that Vpx targets SAMHD1 for degradation in a viral strategy to control cellular deoxynucleotide levels for efficient replication.

FIGURE 1.

SAMHD1 is a nucleotide triphosphohydrolase. A, time course reactions containing 400 nm SAMHD1, 200 μm dGTP, dATP, dTTP, and dCTP, and 5 mm MgCl₂ were performed in triplicate for the indicated times, and products were fractioned by HPLC as described under “Experimental Procedures. ” A chromatogram at each time point from one of the reactions is shown. The indicated positions of elution of the substrates and products were determined in separate injections of each dNTP and nucleoside. B, the dG, dA, dC, and dT products generated at the indicated times from the three reactions in A and from triplicate time course reactions containing only dGTP (dG⁰), only dATP (dA⁰), only dTTP (dT⁰), and only dCTP (dC⁰) were quantified. C, the dG, dA, dT, and dC products from reactions containing 200 or 2000 μm of the indicated nucleotide were quantified, and rates were calculated.

FIGURE 2.

SAMHD1 is a dGTP-regulated triphosphohydrolase. Reactions containing 400 nm SAMHD1, 200 μm dATP, dTTP, or dCTP, 5 mm MgCl₂, and the indicated concentrations of dGTP were performed for 20 min at 25 °C. The reactions were fractioned by HPLC, and the dA (●), dT (▾), and dC (○) products were quantified as described under “Experimental Procedures.” Curved symbols indicate a change in the x-axis scale.