gms | German Medical Science

125. Kongress der Deutschen Gesellschaft für Chirurgie

Deutsche Gesellschaft für Chirurgie

22. - 25.04.2008, Berlin

Systematic Analysis of Combination Effects of HHV-8 genes on NF-κB Activation

Meeting Abstract

  • corresponding author A. Konrad - Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Erlangen, Germany.
  • E. Wies - Institute of Clinical and Molecular Virology, University of Erlangen-Nuremberg, Erlangen, Germany.
  • M. Thurau - Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Erlangen, Germany.
  • G. Sander - Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Erlangen, Germany.
  • E. Naschberger - Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Erlangen, Germany.
  • H. Erfle - MitoCheck Project Group, EMBL, Heidelberg, Germany.
  • F. Neipel - Institute of Clinical and Molecular Virology, University of Erlangen-Nuremberg, Erlangen, Germany.
  • W. Hohenberger - Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Erlangen, Germany.
  • M. Stürzl - Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Erlangen, Germany.

Deutsche Gesellschaft für Chirurgie. 125. Kongress der Deutschen Gesellschaft für Chirurgie. Berlin, 22.-25.04.2008. Düsseldorf: German Medical Science GMS Publishing House; 2008. Doc08dgch9183

Die elektronische Version dieses Artikels ist vollständig und ist verfügbar unter: http://www.egms.de/de/meetings/dgch2008/08dgch340.shtml

Veröffentlicht: 16. April 2008

© 2008 Konrad et al.
Dieser Artikel ist ein Open Access-Artikel und steht unter den Creative Commons Lizenzbedingungen (http://creativecommons.org/licenses/by-nc-nd/3.0/deed.de). Er darf vervielf&aauml;ltigt, verbreitet und &oauml;ffentlich zug&aauml;nglich gemacht werden, vorausgesetzt dass Autor und Quelle genannt werden.


Gliederung

Text

Introduction: Human herpesvirus-8 (HHV-8) is the etiologic agent of Kaposi¡¦s sarcoma (KS), an endothelial cell-derived tumor with pronounced inflammation. Nuclear factor-kappa B (NF-κB) is a key regulatory molecule in inflammation. Using reverse transfection cell arrays (RTCA) as an unbiased systems biology approach the effects on NF-κB-activation of all HHV-8-encoded genes individually and of pairwise combinations of all K- and latent genes were investigated. More than 15.000 transfections were performed.

Materials and methods: All 86 HHV-8 genes were cloned in expression vectors, the resulting plasmids were confirmed in sequence and the expressed proteins were controlled by western blotting and immunofluorescent stainings. NF-κB-activation of all HHV-8-encoded genes individually and of pairwise combinations of all K- and latent genes was investigated through co-transfection with a reporter plasmid expressing GFP under the control of NF-κB in the RTCA manner. Classical transfection experiments were performed with a reporter plasmid expressing firefly luciferase under the control of NF-κB. EMSA was carried out as described previously [1].

Results: The strongest activation of NF-κB was observed in combinations of vFLIP, a known activator of NF-κB, with the latent genes vCYC or LNA-1. Combined activities of these genes were clearly synergistic, could be abrogated by co-expression of a constitutively active IκB-α molecule and were confirmed in classical transfection experiments and EMSA. In addition to the known activator vFLIP, ORF75 was detected as a previously unknown activator of NF-κB. Interestingly NF-κB-activation by ORF75 could not be inhibited by co-expression of a constitutively active IκB-α molecule, suggesting that ORF75 activates NF-κB in the alternative pathway.

Conclusion: Our results indicate that co-operative effects of vFLIP, vCYC and LNA-1, which are all encoded within a tricistronic latency-associated gene locus in the viral genome, and ORF75 are key to the robust NF-κB-activation observed in HHV-8-infected cells.


References

1.
Naschberger et al. Biochem J. 2004.