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Identification of SNF2h, a Chromatin-Remodeling Factor, as a Novel Binding Protein of Vpr of Human Immunodeficiency Virus Type 1

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Abstract

Vpr, an accessory gene of human immunodeficiency virus type 1, encodes a virion-associated nuclear protein that plays an important role in the primary viral infection of resting macrophages. It has a variety of biological functions, including roles in a cell cycle abnormality at G2/M phase, apoptosis, nuclear transfer of preintegration complex, and DNA double-strand breaks (DSBs), some of which depend on its association with the chromatin of the host cells. Given that DSB signals are postulated to be a positive factor in the viral infection, understanding the mode of chromatin recruitment of Vpr is important. Here, we identified SNF2h, a chromatin-remodeling factor, as a novel binding partner of Vpr involved in its chromatin recruitment. When endogenous SNF2h protein was extensively downregulated by SNF2h small interfering RNA (siRNA), the amount of Vpr loaded on chromatin decreased to about 30% of the control level. Biochemical analysis using a mutant Vpr suggested that Vpr binds SNF2h via HFRIG (amino acids 71–75 depicted by single letters) and the Vpr mutant lacking this motif lost the activity to induce DSB-dependent signals. Consistently, Vpr-induced DSBs were attenuated by extensive downregulaion of endogenous SNF2h. Based on these data, we discuss the role of DSB and DSB signals in the viral infection.

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Acknowledgments

We are grateful to Dr. Hiroshi Sasaki (Jikei Medical School) for providing us with 2008 cells. This work was supported in parts by Grants-in-Aid for Research from the Ministry of Health, Labour and Welfare of Japan and for Research on Publicly Essential Drugs and Medical Devices from Japan Health Sciences Foundation Research.

Conflicts of interest

The authors declare no conflicts of interest.

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Authors and Affiliations

  1. Department of Intractable Diseases, National Center for Global Health and Medicine, 1-21-1 Toyama, Shinjuku, Tokyo, 162-8655, Japan

    Daiki Taneichi, Kenta Iijima, Akihiro Doi, Takayoshi Koyama, Yuzuru Minemoto, Mari Shimura & Yukihito Ishizaka

  2. Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Ten-nodai, Tsukuba, 305-8577, Japan

    Daiki Taneichi & Shigeyuki Kano

  3. Department of Pathology, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku, Tokyo, 162-8640, Japan

    Kenzo Tokunaga

  4. Department of Tropical Medicine and Malaria, Research Institute, National Center for Global Health and Medicine, Tokyo, 162-8655, Japan

    Shigeyuki Kano

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  1. Daiki Taneichi
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  2. Kenta Iijima
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  3. Akihiro Doi
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  4. Takayoshi Koyama
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  9. Yukihito Ishizaka
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Correspondence to Yukihito Ishizaka.

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Daiki Taneichi and Kenta Iijima contributed equally to this work.

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Supplementary Figure 1

Effects of SNF2h siRNA on Vpr-induced phosphorylation of ATM. MIT-23 cells were transfected with SNF2h (lower panels) or control (upper panels) siRNAs. DOX was added after 48 h, and an immunohistochemical analysis was carried out after 24 h of the Dox addition. Arrowhead indicates pATM-foci positive cells. Scale bar indicates 10 μm (JPEG 71 kb)

Supplementary Figure 2

rVpr induces focus formation of pATM in resting cells under the downregulated expression of endogenous SNF2h. Serum-starved TIG-3 cells with SNF2h (lower panels) or control (upper panels) siRNAs were further treated with 100 ng/mL of rVpr. Left panel, immunohistochemical analysis of pATM. Arrowhead indicates cells positive for pATM foci (pATM-foci positive cells). Scale bar indicates 10 μm. Right panel, effects of SNF2h siRNA on rVpr-induced focus formation of pATM. Numbers of pATM-foci positive cells were counted three sets of at least 300 cells in each sample and subjected to statistical analysis (a Mann–Whitney test: experiments were performed independently at three times). Mean numbers of pATM-positive cells ± standard deviation were shown. No inhibitory effects of SNF2h siRNA on rVpr-induced DSB were observed (PPT 1143 kb)

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Taneichi, D., Iijima, K., Doi, A. et al. Identification of SNF2h, a Chromatin-Remodeling Factor, as a Novel Binding Protein of Vpr of Human Immunodeficiency Virus Type 1. J Neuroimmune Pharmacol 6, 177–187 (2011). https://doi.org/10.1007/s11481-011-9276-5

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