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Journal of Virology, March 2008, p. 2056-2064, Vol. 82, No. 5
0022-538X/08/$08.00+0     doi:10.1128/JVI.01803-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Dominant-Negative FADD Rescues the In Vivo Fitness of a Cytomegalovirus Lacking an Antiapoptotic Viral Gene

Luka iin-ain,^1, Zsolt Ruzsics,¹ Juergen Podlech,² Ivan Bubi,³ Carine Menard,¹ Stipan Jonji,³ Matthias J. Reddehase,² and Ulrich H. Koszinowski^1*

Max von Pettenkofer Institute, LMU, 80336 Munich, Germany,¹ Institute for Virology, Johannes Gutenberg University, 55131 Mainz, Germany,² Department of Histology and Embryology, Medical Faculty University of Rijeka, 51000 Rijeka, Croatia³

Received 16 August 2007/ Accepted 12 December 2007

Genes that inhibit apoptosis have been described for many DNA viruses. Herpesviruses often contain even more than one gene to control cell death. Apoptosis inhibition by viral genes is postulated to contribute to viral fitness, although a formal proof is pending. To address this question, we studied the mouse cytomegalovirus (MCMV) protein M36, which binds to caspase-8 and blocks death receptor-induced apoptosis. The growth of MCMV recombinants lacking M36 (M36) was attenuated in vitro and in vivo. In vitro, caspase inhibition by zVAD-fmk blocked apoptosis in M36-infected macrophages and rescued the growth of the mutant. In vivo, M36 infection foci in liver tissue contained significantly more apoptotic hepatocytes and Kupffer cells than did revertant virus foci, and apoptosis occurred during the early phase of virus replication prior to virion assembly. To further delineate the mode of M36 function, we replaced the M36 gene with a dominant-negative FADD (FADD^DN) in an MCMV recombinant. FADD^DN was expressed in cells infected with the recombinant and blocked the death-receptor pathway, replacing the antiapoptotic function of M36. Most importantly, FADD^DN rescued M36 virus replication, both in vitro and in vivo. These findings have identified the biological role of M36 and define apoptosis inhibition as a key determinant of viral fitness.

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* Corresponding author. Mailing address: Max von Pettenkofer-Institut, Pettenkoferstrasse 9a, 80336 Munich, Germany. Phone: 49-89-5160-5290. Fax: 49-89-5160-5292. E-mail: koszinowski{at}mvp.uni-muenchen.de

Published ahead of print on 19 December 2007.

Present address: Vaccine and Gene Therapy Institute, Oregon Health and Science University, Beaverton, OR 97006.

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Journal of Virology, March 2008, p. 2056-2064, Vol. 82, No. 5
0022-538X/08/$08.00+0     doi:10.1128/JVI.01803-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

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