The Deoxynucleoside Triphosphate Triphosphohydrolase Activity of SAMHD1 Protein Contributes to the Mitochondrial DNA Depletion Associated with Genetic Deficiency of Deoxyguanosine Kinase.
The dNTP triphosphohydrolase SAMHD1 is a nuclear antiviral host restriction factor limiting HIV-1 infection in macrophages and a major regulator of dNTP concentrations in human cells. In normal human fibroblasts its expression increases during quiescence, contributing to the small dNTP pool sizes of these cells. Down-regulation of SAMHD1 by siRNA expands all four dNTP pools, with dGTP undergoing the largest relative increase. The deoxyguanosine released by SAMHD1 from dGTP can be phosphorylated inside mitochondria by deoxyguanosine kinase (dGK) or degraded in the cytosol by purine nucleoside phosphorylase. Genetic mutations of dGK cause mitochondrial (mt) DNA depletion in noncycling cells and hepato-cerebral mtDNA depletion syndrome in humans. We studied if SAMHD1 and dGK interact in the regulation of the dGTP pool during quiescence employing dGK-mutated skin fibroblasts derived from three unrelated patients. In the presence of SAMHD1 quiescent mutant fibroblasts manifested mt dNTP pool imbalance and mtDNA depletion. When SAMHD1 was silenced by siRNA transfection the composition of the mt dNTP pool approached that of the controls, and mtDNA copy number increased, compensating the depletion to various degrees in the different mutant fibroblasts. Chemical inhibition of purine nucleoside phosphorylase did not improve deoxyguanosine recycling by dGK in WT cells. We conclude that the activity of SAMHD1 contributes to the pathological phenotype of dGK deficiency. Our results prove the importance of SAMHD1 in the regulation of all dNTP pools and suggest that dGK inside mitochondria has the function of recycling the deoxyguanosine derived from endogenous dGTP degraded by SAMHD1 in the nucleus.
From the Department of Biology, University of Padova, 35131 Padova, Italy and.
Elisa
E
Franzolin
Department of Molecular Medicine, University of Padova, 35121 Padova, Italy.
Cristiano
C
Salata
From the Department of Biology, University of Padova, 35131 Padova, Italy and vbianchi@bio.unipd.it.
Vera
V
Bianchi
From the Department of Biology, University of Padova, 35131 Padova, Italy and chiara.rampazzo.1@unipd.it.
Chiara
C
Rampazzo
D004272:DNA, Mitochondrial / Q000235:genetics*; D020868:Gene Silencing; D006801:Humans; D008928:Mitochondria / Q000201:enzymology; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000502:physiology*; D009154:Mutation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics*; D000076106:SAM Domain and HD Domain-Containing Protein 1
D004272:DNA, Mitochondrial; D017853:Phosphotransferases (Alcohol Group Acceptor); C019165:deoxyguanosine kinase; D000076106:SAM Domain and HD Domain-Containing Protein 1; C484280:SAMHD1 protein, human; D020559:Monomeric GTP-Binding Proteins
dGTP pool; gene silencing; mitochondrial DNA (mtDNA); mitochondrial deoxynucleoside salvage; mitochondrial disease; mtDNA precursors; nucleoside/nucleotide metabolism; phosphatase; ribonucleotide reductase