The SAMHD1 dNTP Triphosphohydrolase Is Controlled by a Redox Switch

Abstract

Here we report that SAMHD1 is a major target for redox regulation of nucleotide metabolism and cell cycle control. SAMHD1 is a triphosphate hydrolase, whose function involves regulation of deoxynucleotide triphosphate pools. We demonstrate that the redox state of SAMHD1 regulates its catalytic activity. We have identified three cysteine residues that constitute an intrachain disulfide bond "redox switch" that reversibly inhibits protein tetramerization and catalysis. We show that proliferative signals lead to SAMHD1 oxidation in cells and oxidized SAMHD1 is localized outside of the nucleus. Innovation and Conclusions: SAMHD1 catalytic activity is reversibly regulated by protein oxidation. These data identify a previously unknown mechanism for regulation of nucleotide metabolism by SAMHD1. Antioxid. Redox Signal. 27, 1317-1331.

SAMHD1: Recurring roles in cell cycle, viral restriction, cancer, and innate immunity

“…The sequential activation and assembly of the SAMHD1 tetramer occur at dNTP concentrations multiple orders of magnitude below the K M . Thus, while reported K M values for SAMHD1 vary considerably from 50–350 μM depending on the study, standard Michaelis-Menten kinetic models apply under all the experimental conditions measured (55, 57, 73, 78, 81, 82). Given the steady-state conditions of an activated SAMHD1 tetramer, the turnover rate for each substrate appears to depend solely on its particular affinity for the active site and its intracellular concentration.…”

“…A recent study demonstrated that SAMHD1 catalytic activity is regulated by redox signaling (82). Redox signaling takes the form of reactive oxygen species (ROS) that are generated by cellular oxidases in response to growth factors binding and activating cellular receptors (130).…”

“…These ROS act as secondary messengers that can covalently, but reversibly, interact with an enzyme to modify its tertiary structure and subsequent activity(131). SAMHD1 is catalytically inactivated when treated with the oxidizing agent H 2 O 2 in a dose dependent but reversible manner (82). Oxidation of SAMHD1 was demonstrated to inhibit tetramerization, but this deficiency was ablated in SAMHD1 mutants containing a C522A mutation.…”

“…The redox sensitivity of SAMHD1 was recapitulated in cells during experiments in which SAMHD1 was demonstrated to be oxidized in response to proliferative signals (82). It was proposed that cells oxidize SAMHD1 in response to proliferative signaling in order to accumulate the dNTPs necessary for DNA replication.…”

Mapping disulfide bonds from sub-micrograms of purified proteins or micrograms of complex protein mixtures

“…In 2015, our laboratory developed a sensitive and efficient method for mapping protein disulfide bonds from simple or complex samples (Lu et al . in Nat Methods 12:329, 2015). This method is based on liquid chromatography–mass spectrometry (LC–MS) and a powerful data analysis software tool named pLink.…”