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Human Immunodeficiency Virus Type 1 Vpr Induces DNA Replication Stress In Vitro and In Vivo

Erik S. Zimmerman,^1, Michael P. Sherman,^2, Jana L. Blackett,¹ Jason A. Neidleman,² Christophe Kreis,² Pamela Mundt,² Samuel A. Williams,² Maria Warmerdam,² James Kahn,³ Frederick M. Hecht,³ Robert M. Grant,² Carlos M. C. de Noronha,² Andrew S. Weyrich,⁴ Warner C. Greene,² and Vicente Planelles^1*

Division of Cellular Biology and Immunology, Department of Pathology, University of Utah School of Medicine, Salt Lake City, Utah 84132,¹ Gladstone Institute of Virology and Immunology, San Francisco, California 94158,² Positive Health Program, San Francisco General Hospital, San Francisco, California 94110,³ Program in Human Molecular Biology and Genetics, Department of Internal Medicine, University of Utah School of Medicine, Salt Lake City, Utah 84132⁴

Received 9 June 2006/ Accepted 10 August 2006

The human immunodeficiency virus type 1 (HIV-1) viral protein R (Vpr) causes cell cycle arrest in G₂. Vpr-expressing cells display the hallmarks of certain forms of DNA damage, specifically activation of the ataxia telangiectasia mutated and Rad3-related kinase, ATR. However, evidence that Vpr function is relevant in vivo or in the context of viral infection is still lacking. In the present study, we demonstrate that HIV-1 infection of primary, human CD4⁺ lymphocytes causes G₂ arrest in a Vpr-dependent manner and that this response requires ATR, as shown by RNA interference. The event leading to ATR activation in CD4⁺ lymphocytes is the accumulation of replication protein A in nuclear foci, an indication that Vpr likely induces stalling of replication forks. Primary macrophages are refractory to ATR activation by Vpr, a finding that is consistent with the lack of detectable ATR, Rad17, and Chk1 protein expression in these nondividing cells. These observations begin to explain the remarkable resilience of macrophages to HIV-1-induced cytopathicity. To study the in vivo consequences of Vpr function, we isolated CD4⁺ lymphocytes from HIV-1-infected individuals and interrogated the cell cycle status of anti-p24^Gag-immunoreactive cells. We report that infected cells in vivo display an aberrant cell cycle profile whereby a majority of cells have a 4N DNA content, consistent with the onset of G₂ arrest.

Published ahead of print on 6 September 2006.

These authors contributed equally.

This article has been cited by other articles:

• DeHart, J. L., Planelles, V. (2008). Human Immunodeficiency Virus Type 1 Vpr Links Proteasomal Degradation and Checkpoint Activation. J. Virol. 82: 1066-1072 [Full Text]  
• Schrofelbauer, B., Hakata, Y., Landau, N. R. (2007). HIV-1 Vpr function is mediated by interaction with the damage-specific DNA-binding protein DDB1. Proc. Natl. Acad. Sci. USA 104: 4130-4135 [Abstract] [Full Text]