SUMMARY DNA double-strand break (DSB) repair by homologous recombination (HR) is initiated by CtIP/MRN-mediated DNA end resection to maintain genome integrity. SAMHD1 is a dNTP triphosphohydrolase, which restricts HIV-1 infection, and mutations are associated with Aicardi-Goutières syndrome and cancer. We show that SAMHD1 has a dNTPase-independent function in promoting DNA end resection to facilitate DSB repair by HR. SAMHD1 deficiency or Vpx-mediated degradation causes hypersensitivity to DSB-inducing agents, and SAMHD1 is recruited to DSBs. SAMHD1 complexes with CtIP via a conserved carboxyl-terminal domain and recruits CtIP to DSBs to facilitate end resection and HR. Significantly, a cancer-associated mutant with impaired CtIP interaction but not dNTPase-inactive SAMHD1 fails to rescue the end resection impairment of SAMHD1 depletion. Our findings define a dNTPase-independent function for SAMHD1 in HR-mediated DSB repair by facilitating CtIP accrual to promote DNA end resection, providing insight into how SAMHD1 promotes genome integrity and prevents disease, including cancer.

SAMHD1 is a newly identified anti-HIV host factor that has a dNTP triphosphohydrolase activity and depletes intracellular dNTP pools in non-dividing myeloid cells. Since DNA viruses utilize cellular dNTPs, we investigated whether SAMHD1 limits the replication of DNA viruses in non-dividing myeloid target cells. Indeed, two double stranded DNA viruses, vaccinia and herpes simplex virus type 1, are subject to SAMHD1 restriction in non-dividing target cells in a dNTP dependent manner. Using a thymidine kinase deficient strain of vaccinia virus, we demonstrate a greater restriction of viral replication in non-dividing cells expressing SAMHD1. Therefore, this study suggests that SAMHD1 is a potential innate anti-viral player that suppresses the replication of a wide range of DNA viruses, as well as retroviruses, which infect non-dividing myeloid cells.

SAMHD1 limits HIV-1 infection in non-dividing myeloid cells by decreasing intracellular dNTP pools. HIV-1 restriction by SAMHD1 in these cells likely prevents activation of antiviral immune responses and modulates viral pathogenesis, thus highlighting a critical role of SAMHD1 in HIV-1 physiopathology. Here, we explored the function of SAMHD1 in regulating cell proliferation, cell cycle progression and apoptosis in monocytic THP-1 cells. Using the CRISPR/Cas9 technology, we generated THP-1 cells with stable SAMHD1 knockout. We found that silencing of SAMHD1 in cycling cells stimulates cell proliferation, redistributes cell cycle population in the G1/G0 phase and reduces apoptosis. These alterations correlated with increased dNTP levels and more efficient HIV-1 infection in dividing SAMHD1 knockout cells relative to control. Our results suggest that SAMHD1, through its dNTPase activity, affects cell proliferation, cell cycle distribution and apoptosis, and emphasize a key role of SAMHD1 in the interplay between cell cycle regulation and HIV-1 infection.

We previously reported that combining a phosphoinositide 3-kinase (PI3K) inhibitor with a poly-ADP Rib polymerase (PARP)-inhibitor enhanced DNA damage and cell death in breast cancers that have genetic aberrations in BRCA1 and TP53. Here, we show that enhanced DNA damage induced by PI3K inhibitors in this mutational background is a consequence of impaired production of nucleotides needed for DNA synthesis and DNA repair. Inhibition of PI3K causes a reduction in all four nucleotide triphosphates, whereas inhibition of the protein kinase AKT is less effective than inhibition of PI3K in suppressing nucleotide synthesis and inducing DNA damage. Carbon flux studies reveal that PI3K inhibition disproportionately affects the nonoxidative pentose phosphate pathway that delivers Rib-5-phosphate required for base ribosylation. In vivo in a mouse model of BRCA1-linked triple-negative breast cancer (K14-Cre BRCA1), the PI3K inhibitor BKM120 led to a precipitous drop in DNA synthesis within 8 h of drug treatment, whereas DNA synthesis in normal tissues was less affected. In this mouse model, combined PI3K and PARP inhibition was superior to either agent alone to induce durable remissions of established tumors. T riple-negative breast cancers, including BRCA1-linked breast cancers, frequently show activation of the PI3K pathway as a result of overexpression of epidermal growth factor receptor (EGFR) (1, 2) or insulin-like growth factor 1 receptor (IGFR) (3), and inhibition of the EGFR (1, 2, 4) and/or the PI3K/Nrf2 axis (5) prevents cancers arising from BRCA1 mutant mammary epithelial cells (MECs). In addition, activating mutations of PIK3CA, or loss of the inhibitory lipid phosphatases PTEN (phosphatase and tensin homolog) and INPP4B (inositol polyphosphate 4-phosphatase type II) (6, 7), suggest that the PI3K pathway is contributing to tumor growth and survival. Aside from their role in regulating the homeostasis of phospho-inositides, PTEN and INPP4B may have independent roles in DNA damage repair. A role for PTEN in the maintenance of genomic stability was identified (8); more recently, INPP4B was found to directly interact with BRCA1 and the serine/threonine protein kinase ATR, and its loss destabilizes these DNA damage repair complexes, effectively sensitizing INPP4B-deficient cells to poly-ADP Rib polymerase (PARP) inhibition (9).Despite the high incidence of predisposing lesions in the PI3K pathway, limited clinical activity has been observed with PI3K inhibitors as single-agent treatment in endocrine-resistant breast cancer, which may reflect bypass of PI3K-dependent mitogenic signaling by alternative signaling pathways such as the MAPK pathway. Therefore, concurrent inhibition of parallel and compensatory signaling networks to overcome resistance to PI3K inhibition is being investigated in clinical studies. This approach, however, carries the risk of overlapping toxicities of the targeted agents without sufficient efficacy because tumor cells may have greater plasticity for redundant signaling than normal tissues.Mul...