Cyclin D3-dependent control of the dNTP pool and HIV-1 replication in human macrophages

Abstract

Cyclins control the activation of cyclin-dependent kinases (CDK), which in turn, control the cell cycle and cell division. Intracellular availability of deoxynucleotides (dNTP) plays a fundamental role in cell cycle progression. SAM domain and HD domain-containing protein 1 (SAMHD1) degrades nucleotide triphosphates and controls the size of the dNTP pool. SAMHD1 activity appears to be controlled by CDK. Here, we show that knockdown of cyclin D3 a partner of CDK6 and E2 a partner of CDK2 had a major impact in SAMHD1 phosphorylation and inactivation and led to decreased dNTP levels and inhibition of HIV-1 at the reverse transcription step in primary human macrophages. The effect of cyclin D3 RNA interference was lost after degradation of SAMHD1 by HIV-2 Vpx, demonstrating the specificity of the mechanism. Cyclin D3 inhibition correlated with decreased activation of CDK2. Our results confirm the fundamental role of the CDK6-cyclin D3 pair in controlling CDK2-dependent SAMHD1 phosphorylation and dNTP pool in primary macrophages.

SAMHD1: At the Crossroads of Cell Proliferation, Immune Responses, and Virus Restriction

“…Knockdown of CDK6 or CDK2, but not CDK1, in proliferating macrophages blocked SAMHD1 phosphorylation and restricted HIV-1 infection [32]. Moreover, knockdown of cyclin D3, a cyclin partner controlling CDK6 and a selective inhibitor of CDK6, potently blocked SAMHD1 phosphorylation, reduced dNTP levels, and inhibited HIV-1 replication at the reverse transcriptase step [48,49]. The observation that p21, a natural inhibitor of CDK2, affected …”

Subversion of Cell Cycle Regulatory Mechanisms by HIV

“…This conundrum was resolved when it was discovered that cyclin-dependent kinases (CDKs) activated during the cell cycle of proliferating cells can inactivate SAMHD1 through phosphorylation of threonine 592 (T592) (White et al, 2013; Cribier et al, 2013). The mechanism whereby this phosphorylation inactivates SAMDH1 is presently unclear, as there are conflicting reports on the effects on phosphohydrolase activity (White et al, 2013; Yan et al, 2015; Ruiz et al, 2015). Additional studies are required to clarify to what extent T592 phosphorylation inhibits SAMHD1 phosphohydrolase and nuclease activity, and the relative importance of each activity for anti-HIV-1 activity.…”

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

“…The precise effects of phosphorylation on SAMHD1 activity are still under debate, however. Some evidence suggests phosphorylation negatively modulates SAMHD1 tetramerization and dNTPase activity (81, 99, 127), and this diminished dNTPase capacity is responsible for increased intracellular dNTP pools (93, 120, 121, 128). Other studies find limited or no effect of phosphorylation of SAMHD1 on catalytic activity(81, 108, 116, 117, 129), oligomerization equilibrium and allosteric activation(77, 129), or nucleic acid binding (83).…”