Abstract
ABSTRACTSAMHD1 is the major catabolic enzyme regulating the intracellular concentrations of DNA precursors (dNTPs). The S-phase kinase CDK2-cyclinA phosphorylates SAMHD1 at Thr-592. How this modification affects SAMHD1 function is highly debated. We investigated the role of endogenous SAMHD1 phosphorylation during the cell cycle. Thr-592 phosphorylation occurs first at the G1/S border and is removed during mitotic exit parallel with Thr-phosphorylations of most CDK1 targets. Differential sensitivity to the phosphatase inhibitor okadaic acid suggested different involvement of the PP1 and PP2 families dependent upon the time of the cell cycle. SAMHD1 turn-over indicates that Thr-592 phosphorylation does not cause rapid protein degradation. Furthermore, SAMHD1 influenced the size of the four dNTP pools independently of its phosphorylation. Our findings reveal that SAMHD1 is active during the entire cell cycle and performs an important regulatory role during S-phase by contributing with ribonucleotide reductase to maintain dNTP pool balance for proper DNA replication.
Highlights
The Sterile α motif domain and histidine-aspartate domain-containing protein 1 (SAMHD1) is a nuclear triphosphohydrolase that cleaves all four deoxynucleoside triphosphates to deoxynucleosides and inorganic triphosphate
The absence of SAMHD1 in THP-1 KO cells leaves the expression of ribonucleotide reductase (RNR) subunits unaffected and causes a strong increase in deoxynucleoside triphosphates (dNTPs) pools in all phases of the cell cycle
It is still controversial if phosphorylation at threonine 592 (T592) impairs SAMHD1 dNTPase activity, since in vitro data obtained with purified SAMHD1 variants do not match the effects of the same mutants on the dNTP pools of transfected cells [15,20,21,22]
Summary
The Sterile α motif domain and histidine-aspartate domain-containing protein 1 (SAMHD1) is a nuclear triphosphohydrolase that cleaves all four deoxynucleoside triphosphates (dNTPs) to deoxynucleosides and inorganic triphosphate. The catalytic activity is regulated by nucleoside triphosphate binding at two allosteric sites, which induces the formation of a stable tetramer [1]. Later data attributed the nuclease activity to contaminants co-purifying with SAMHD1 and the question of SAMHD1 harboring multiple functions is still debated [5]. SAMHD1 is expressed at variable levels in most human tissues, especially in immune cells. It has been intensively investigated as a host restriction factor that, in quiescent/ differentiated cells, limits HIV-1 and other viral infections by lowering cellular dNTP concentrations under a threshold critical for the synthesis of viral DNA [6]
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