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Journal of Virology, March 2009, p. 2563-2574, Vol. 83, No. 6
0022-538X/09/$08.00+0 doi:10.1128/JVI.01512-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
A Systems Biology Approach To Identify the Combination Effects of Human Herpesvirus 8 Genes on NF-
B Activation
Andreas Konrad,1
Effi Wies,2
Mathias Thurau,1
Gaby Marquardt,1
Elisabeth Naschberger,1
Sonja Hentschel,3
Ramona Jochmann,1
Thomas F. Schulz,4
Holger Erfle,5
Benedikt Brors,6
Berthold Lausen,7
Frank Neipel,2 and
Michael Stürzl1*
Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Schwabachanlage 10, D-91054 Erlangen, Germany,1 Institute of Clinical and Molecular Virology, University of Erlangen-Nuremberg, Schlossgarten 4, D-91054 Erlangen, Germany,2 Department of Virus-Induced Vasculopathy, Helmholtz-Center Munich, Ingolstädter Landstrasse 1, D-85764 Neuherberg, Germany,3 Medical School Hannover, Department of Virology, Carl-Neuberg-Strasse 1, D-30625 Hannover, Germany,4 VIROQUANT-Cell Networks RNAi Screening Facility, BQ 0015, BIOQUANT-Zentrum, Ruprecht-Karls-Universität Heidelberg, Im Neuenheimer Feld 267, D-69120 Heidelberg, Germany,5 Intelligent Bioinformatics Systems, iBios, DKFZ Heidelberg, Im Neuenheimer Feld 580, D-69120 Heidelberg, Germany,6 Institute of Medical Informatics, Biometry, and Epidemiology, University of Erlangen-Nuremberg, Waldstrasse 6, D-91054 Erlangen, Germany7
Received 18 July 2008/ Accepted 1 January 2009
Human herpesvirus 8 (HHV-8) is the etiologic agent of Kaposi's sarcoma and primary effusion lymphoma. Activation of the cellular transcription factor nuclear factor-kappa B (NF-
B) is essential for latent persistence of HHV-8, survival of HHV-8-infected cells, and disease progression. We used reverse-transfected cell microarrays (RTCM) as an unbiased systems biology approach to systematically analyze the effects of HHV-8 genes on the NF-B signaling pathway. All HHV-8 genes individually (n = 86) and, additionally, all K and latent genes in pairwise combinations (n = 231) were investigated. Statistical analyses of more than 14,000 transfections identified ORF75 as a novel and confirmed K13 as a known HHV-8 activator of NF-B. K13 and ORF75 showed cooperative NF-B activation. Small interfering RNA-mediated knockdown of ORF75 expression demonstrated that this gene contributes significantly to NF-B activation in HHV-8-infected cells. Furthermore, our approach confirmed K10.5 as an NF-B inhibitor and newly identified K1 as an inhibitor of both K13- and ORF75-mediated NF-B activation. All results obtained with RTCM were confirmed with classical transfection experiments. Our work describes the first successful application of RTCM for the systematic analysis of pathofunctions of genes of an infectious agent. With this approach, ORF75 and K1 were identified as novel HHV-8 regulatory molecules on the NF-B signal transduction pathway. The genes identified may be involved in fine-tuning of the balance between latency and lytic replication, since this depends critically on the state of NF-B activity.
* Corresponding author. Mailing address: University of Erlangen-Nuremberg, Department of Surgery, Division of Molecular and Experimental Surgery, Schwabachanlage 10, D-91054 Erlangen, Germany. Phone: 49-9131-85-33109. Fax: 49-9131-85-32077. E-mail: michael.stuerzl{at}uk-erlangen.de
Published ahead of print on 7 January 2009.
Journal of Virology, March 2009, p. 2563-2574, Vol. 83, No. 6
0022-538X/09/$08.00+0 doi:10.1128/JVI.01512-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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