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A viral kinase counteracts in vivo restriction of murine cytomegalovirus by SAMHD1

Nature Microbiology volume 4, pages 2273–2284 (2019)Cite this article

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Abstract

The deoxynucleotide triphosphate (dNTP) hydrolase SAMHD1 inhibits retroviruses in non-dividing myeloid cells. Although antiviral activity towards DNA viruses has also been demonstrated, the role of SAMHD1 during cytomegalovirus (CMV) infection remains unclear. To determine the impact of SAMHD1 on the replication of CMV, we used murine CMV (MCMV) to infect a previously established SAMHD1 knockout mouse model and found that SAMHD1 inhibits the replication of MCMV in vivo. By comparing the replication of MCMV in vitro in myeloid cells and fibroblasts from SAMHD1-knockout and control mice, we found that the viral kinase M97 counteracts SAMHD1 after infection by phosphorylating the regulatory residue threonine 603. The phosphorylation of SAMHD1 in infected cells correlated with a reduced level of dNTP hydrolase activity and the loss of viral restriction. Together, we demonstrate that SAMHD1 acts as a restriction factor in vivo and we identify the M97-mediated phosphorylation of SAMHD1 as a previously undescribed viral countermeasure.

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Fig. 1: MCMV replication is enhanced in SAMHD1 KO mice in vivo.
Fig. 2: SAMHD1 does not affect infectivity in vitro and is phosphorylated after MCMV infection.
Fig. 3: WT M97, but not the KD mutant, induces phosphorylation of SAMHD1.
Fig. 4: Increased dNTP levels after infection with MCMV WT but not M97 KD MCMV.
Fig. 5: Replication of M97-defective MCMV is sensitive to SAMHD1-mediated restriction in vitro and in vivo.
Fig. 6: M97-induced phosphorylation at T603 counteracts SAMHD1 restriction.

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Data availability

The data that support the findings of the study are available from the corresponding author on reasonable request.

References

  1. Franzolin, E. et al. The deoxynucleotide triphosphohydrolase SAMHD1 is a major regulator of DNA precursor pools in mammalian cells. Proc. Natl Acad. Sci. USA 110, 14272–14277 (2013).

    Article  CAS  Google Scholar 

  2. Goldstone, D. C. et al. HIV-1 restriction factor SAMHD1 is a deoxynucleoside triphosphate triphosphohydrolase. Nature 480, 379–382 (2011).

    Article  CAS  Google Scholar 

  3. Powell, R. D., Holland, P. J., Hollis, T. & Perrino, F. W. Aicardi-Goutieres syndrome gene and HIV-1 restriction factor SAMHD1 is a dGTP-regulated deoxynucleotide triphosphohydrolase. J. Biol. Chem. 286, 43596–43600 (2011).

    Article  CAS  Google Scholar 

  4. Hrecka, K. et al. Vpx relieves inhibition of HIV-1 infection of macrophages mediated by the SAMHD1 protein. Nature 474, 658–661 (2011).

    Article  CAS  Google Scholar 

  5. Laguette, N. et al. SAMHD1 is the dendritic- and myeloid-cell-specific HIV-1 restriction factor counteracted by Vpx. Nature 474, 654–657 (2011).

    Article  CAS  Google Scholar 

  6. Gramberg, T. et al. Restriction of diverse retroviruses by SAMHD1. Retrovirology 10, 26 (2013).

    Article  CAS  Google Scholar 

  7. Kim, E. T., White, T. E., Brandariz-Nunez, A., Diaz-Griffero, F. & Weitzman, M. D. SAMHD1 restricts herpes simplex virus 1 in macrophages by limiting DNA replication. J. Virol. 87, 12949–12956 (2013).

    Article  CAS  Google Scholar 

  8. Hollenbaugh, J. A. et al. Host factor SAMHD1 restricts DNA viruses in non-dividing myeloid cells. PLoS Pathog. 9, e1003481 (2013).

    Article  CAS  Google Scholar 

  9. Lahouassa, H. et al. SAMHD1 restricts the replication of human immunodeficiency virus type 1 by depleting the intracellular pool of deoxynucleoside triphosphates. Nat. Immunol. 13, 223–228 (2012).

    Article  CAS  Google Scholar 

  10. Cribier, A., Descours, B., Valadao, A. L., Laguette, N. & Benkirane, M. Phosphorylation of SAMHD1 by cyclin A2/CDK1 regulates its restriction activity toward HIV-1. Cell Rep. 3, 1036–1043 (2013).

    Article  CAS  Google Scholar 

  11. White, T. E. et al. The retroviral restriction ability of SAMHD1, but not its deoxynucleotide triphosphohydrolase activity, is regulated by phosphorylation. Cell Host Microbe 13, 441–451 (2013).

    Article  CAS  Google Scholar 

  12. Tramentozzi, E. et al. The dNTP triphosphohydrolase activity of SAMHD1 persists during S-phase when the enzyme is phosphorylated at T592. Cell Cycle 17, 1202–1214 (2018).

    Article  Google Scholar 

  13. Herrmann, A. et al. The SAMHD1-mediated block of LINE-1 retroelements is regulated by phosphorylation. Mob. DNA 9, 11 (2018).

    Article  Google Scholar 

  14. Behrendt, R. et al. Mouse SAMHD1 has antiretroviral activity and suppresses a spontaneous cell-intrinsic antiviral response. Cell Rep. 4, 689–696 (2013).

    Article  CAS  Google Scholar 

  15. Rehwinkel, J. et al. SAMHD1-dependent retroviral control and escape in mice. EMBO J. 32, 2454–2462 (2013).

    Article  CAS  Google Scholar 

  16. Wittmann, S. et al. Phosphorylation of murine SAMHD1 regulates its antiretroviral activity. Retrovirology 12, 103 (2015).

    Article  Google Scholar 

  17. Prichard, M. N. Function of human cytomegalovirus UL97 kinase in viral infection and its inhibition by maribavir. Rev. Med. Virol. 19, 215–229 (2009).

    Article  CAS  Google Scholar 

  18. Marschall, M., Feichtinger, S. & Milbradt, J. Regulatory roles of protein kinases in cytomegalovirus replication. Adv. Virus Res. 80, 69–101 (2011).

    Article  CAS  Google Scholar 

  19. Wagner, M. et al. Comparison between human cytomegalovirus pUL97 and murine cytomegalovirus (MCMV) pM97 expressed by MCMV and vaccinia virus: pM97 does not confer ganciclovir sensitivity. J. Virol. 74, 10729–10736 (2000).

    Article  CAS  Google Scholar 

  20. Rawlinson, W. D. et al. The murine cytomegalovirus (MCMV) homolog of the HCMV phosphotransferase (UL97(pk)) gene. Virology 233, 358–363 (1997).

    Article  CAS  Google Scholar 

  21. Klenovsek, K. et al. Protection from CMV infection in immunodeficient hosts by adoptive transfer of memory B cells. Blood 110, 3472–3479 (2007).

    Article  CAS  Google Scholar 

  22. Jordan, S. et al. Virus progeny of murine cytomegalovirus bacterial artificial chromosome pSM3fr show reduced growth in salivary glands due to a fixed mutation of MCK-2. J. Virol. 85, 10346–10353 (2011).

    Article  CAS  Google Scholar 

  23. Sinzger, C., Plachter, B., Grefte, A., The, T. H. & Jahn, G. Tissue macrophages are infected by human cytomegalovirus in vivo. J. Infect. Dis. 173, 240–245 (1996).

    Article  CAS  Google Scholar 

  24. Ibanez, C. E., Schrier, R., Ghazal, P., Wiley, C. & Nelson, J. A. Human cytomegalovirus productively infects primary differentiated macrophages. J. Virol. 65, 6581–6588 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Stoddart, C. A. et al. Peripheral blood mononuclear phagocytes mediate dissemination of murine cytomegalovirus. J. Virol. 68, 6243–6253 (1994).

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Yamaguchi, T., Shinagawa, Y. & Pollard, R. B. Relationship between the production of murine cytomegalovirus and interferon in macrophages. J. Gen. Virol. 69, 2961–2971 (1988).

    Article  CAS  Google Scholar 

  27. Chong, K. T., Gresser, I. & Mims, C. A. Interferon as a defence mechanism in mouse cytomegalovirus infection. J. Gen. Virol. 64, 461–464 (1983).

    Article  CAS  Google Scholar 

  28. Welbourn, S., Dutta, S. M., Semmes, O. J. & Strebel, K. Restriction of virus infection but not catalytic dNTPase activity are regulated by phosphorylation of SAMHD1. J. Virol. 87, 11516–11524 (2013).

    Article  CAS  Google Scholar 

  29. Detje, C. N. et al. Local type I IFN receptor signaling protects against virus spread within the central nervous system. J. Immunol. 182, 2297–2304 (2009).

    Article  CAS  Google Scholar 

  30. Kamphuis, E., Junt, T., Waibler, Z., Forster, R. & Kalinke, U. Type I interferons directly regulate lymphocyte recirculation and cause transient blood lymphopenia. Blood 108, 3253–3261 (2006).

    Article  CAS  Google Scholar 

  31. Sell, S. et al. Control of murine cytomegalovirus infection by γδ T cells. PLoS Pathog. 11, e1004481 (2015).

    Article  Google Scholar 

  32. Tischer, B. K., Smith, G. A. & Osterrieder, N. En passant mutagenesis: a two step markerless red recombination system. Methods Mol. Biol. 634, 421–430 (2010).

    Article  CAS  Google Scholar 

  33. Zufferey, R., Nagy, D., Mandel, R. J., Naldini, L. & Trono, D. Multiply attenuated lentiviral vector achieves efficient gene delivery in vivo. Nat. Biotechnol. 15, 871–875 (1997).

    Article  CAS  Google Scholar 

  34. Zhang, R., Bloch, N., Nguyen, L. A., Kim, B. & Landau, N. R. SAMHD1 restricts HIV-1 replication and regulates interferon production in mouse myeloid cells. PLoS ONE 9, e89558 (2014).

    Article  Google Scholar 

  35. Reed, L. J. & Muench, H. A simple method of estimating 50 percent endpoints (TCID50). Am. J. Hyg. 23, 493–497 (1938).

    Google Scholar 

  36. Thomas, D., Herold, N., Keppler, O. T., Geisslinger, G. & Ferreiros, N. Quantitation of endogenous nucleoside triphosphates and nucleosides in human cells by liquid chromatography tandem mass spectrometry. Anal. Bioanal. Chem. 407, 3693–3704 (2015).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank M. Marschall, M. Mach, B. Kropf and M. Thomas from the Institute of Virology in Erlangen, D. Dudziak and C. Lehmann from the Dermatology Department in Erlangen, and R. Behrendt and D. Lindemann from TU Dresden for their expertise and the sharing of methods and reagents. T.G. and J.D. were funded by the German research foundation (GR3355/3-1) and the J. und F. Marohn Stiftung. J.M. and M.K. were supported by the German research foundation (MI2143/2-1). T.H.W. was supported by grant number SFB643 TP C09. N.F. and D.T. were supported by the LOEWE program from the state of Hesse (Translational Medicine and Pharmacology). Supplementary Fig. 1 is based on artwork provided by servier.com.

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Authors and Affiliations

  1. Institute of Clinical and Molecular Virology, Friedrich-Alexander University Erlangen-Nürnberg, Erlangen, Germany

    Janina Deutschmann, Melissa Kießling, Sabine Wittmann, Alexandra Herrmann, Jens Milbradt & Thomas Gramberg

  2. Chair of Genetics, Department of Biology, Nikolaus-Fiebiger-Center for Molecular Medicine, Friedrich-Alexander-University Erlangen-Nürnberg, Erlangen, Germany

    Andrea Schneider & Thomas H. Winkler

  3. Laboratory of Molecular Pediatrics, Department of Pediatric Oncology, Hematology and Stem Cell Transplantation, Charité Universitätsmedizin, Berlin, Germany

    Iris Gruska, Barbara Vetter & Lüder Wiebusch

  4. Pharmazentrum Frankfurt/ZAFES, Institute of Clinical Pharmacology, Goethe University, Frankfurt, Germany

    Dominique Thomas & Nerea Ferreirós

  5. Institute for Medical Virology, University Hospital Tübingen, Tübingen, Germany

    Michael Schindler

  6. Project Group Translational Medicine and Pharmacology TMP, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Frankfurt, Germany

    Nerea Ferreirós

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Contributions

T.G., J.D., T.H.W., M.S. and L.W. designed the project. J.D. generated cell lines, viral amplification, quantification and kinetics. J.D. and A.S. performed the mouse infection experiments. J.D., M.K., J.M. and I.G. analysed SAMHD1 phosphorylation in vitro. B.V., A.H. and S.W. performed the cloning and mouse breeding. J.D., D.T. and N.F. quantified dNTPs by HPLC and M.S., J.D. and T.G. wrote the manuscript.

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Correspondence to Thomas Gramberg.

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Supplementary Figs. 1–6, Supplementary references, raw immunoblot data.

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Deutschmann, J., Schneider, A., Gruska, I. et al. A viral kinase counteracts in vivo restriction of murine cytomegalovirus by SAMHD1. Nat Microbiol 4, 2273–2284 (2019). https://doi.org/10.1038/s41564-019-0529-z

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