SFB796 - Sub project B1

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STAT-Activation in T-cell growth transformation

 

Project summary

Reprogramming of human cells by persistent DNA viruses depends on compatible interactions between virus-encoded and host factors to facilitate host cell survival or expansion, genome (co)replication and persistence. In the first funding period, STAT5 activation was studied in virus transformed T-cells. We could demonstrate a direct interaction between the viral effector Tip and STAT5 signalling proteins, identify the responsible domain, and show that it is required for T-cell growth transformation. Furthermore, the viral tegument protein Orf3 was found to be responsible for selective degradation of SP100, conferring an unprecedented function in evading the host cell ND10 resistance complex.

In the second phase of the CRC796, we would like to understand the key role of STAT factor driven intracellular signalling in virus-mediated T-cell transformation. We hypothesize that STAT activation by the viral oncoprotein Tip results in a stable, epigenetically fixed host cell reprogramming; we are specifically interested in the epigenetic chromatin signature of the Tip-driven STAT activation on the host cell genome. The importance of this interaction will be tested primarily by designing respective recombinant viruses with Tip mutations for studies in transformed cells, and by lentiviral shRNA knockdown of STAT. The initial phase of host cell reprogramming by Tip will be studied by electroporation of Tip encoding mRNA and marker viruses. Furthermore, interaction partners of Tip/STAT as well as Orf3 will be searched and identified by proteomics (co-immunoprecipitation coupled to mass spectrometry). The Orf3 interaction will be narrowed down to functional residues, to separate tegument function and ND10 modulation. Orf3 recombinant viruses with preserved tegument function but impaired in ND10 modulation will be used to reveal the consequences of the ND10 directed effector function of Orf3 on the modification status of viral chromatin, and thereby the establishment of viral latency in transformed cell lines. This will lead to a better understanding of host cell epigenetic changes driven by viral effectors, and the functional consequences for both the host cell and the virus.

 

Fig. 1:   STAT transcription factors are key molecules for gene expression after growth factor or cytokine stimulus. Activation of STAT pathways by mutated Janus tyrosine kinases can be observed in tumors, e.g. JAK1 in T cell acute leukemia (T-ALL). The viral effector Tip activates different STAT-transcription factors via the lymphocyte tyrosine kinase Lck. Tip thereby imitates a cytokine/growth factor signal; this results in proliferation and altered differentiation of the infected cell.

 

Fig. 2:   SP100 degradation by the viral ORF3 encoded FGARAT.

 

Project relevant publications

  • Proff J, Walterskirchen C, Brey C, Geyeregger R, Full F, Ensser A, Lehner M, Holter W.   (2016).   Cytomegalovirus-infected cells resist T cell mediated killing in an HLA-recognition independent manner.   Frontiers in Microbiology   7. doi: 10.3389/fmicb.2016.00844.

  • Brulois K, Wong LY, Lee HR, Sivadas P, Ensser A, Feng P, Gao SJ, Toth Z, Jung JU.   (2015).   Association of Kaposi's Sarcoma-Associated Herpesvirus ORF31 with ORF34 and ORF24 Is Critical for Late Gene Expression.   J Virol.   89: 6149-54.

  • Full F, Jungnickl D, Reuter N, Bogner E, Brulois K, Scholz B, Stürzl M, Myoung J, Jung JU, Stamminger T., Ensser A.   (2014).   Kaposi's sarcoma associated herpesvirus tegument protein ORF75 is essential for viral lytic replication and plays a critical role in the antagonization of ND10-instituted intrinsic immunity.   PLoS Pathog.   10: e1003863.

  • Vogel B, Tennert K, Full F, Ensser A.   (2014).   Growth Transformation of Human Natural Killer cells by a Gammaherpesvirus.   Leukemia   28: 192-5. doi:10.1038/leu.2013.188.

  • Brulois K, Toth Z, Wong LY, Feng P, Gao SJ, Ensser A, Jung JU.   (2014).   Kaposi's Sarcoma-Associated Herpesvirus K3 and K5 Ubiquitin E3 Ligases Have Stage-Specific Immune Evasion Roles during Lytic Replication.   J Virol.   88: 9335-49.

  • Wong LY, Brulois K, Toth Z, Inn K-S, Lee SH, O'Brien K, Lee H, Gao SJ, Cesarman E, Ensser A, Jung JU.   (2013).   KSHV K4.2 immediate early gene product regulates immunoglobulin secretion and calcium homeostasis by interacting with and inhibiting pERP1.   J Virol.   87: 12069-12079.

  • Ott L, Scholz B, Höller M, Hasselt K, Ensser A, Burkovski A.   (2013).   Induction of the NFκ-B signal transduction pathway in response to Corynebacterium diphtheriae infection.   Microbiology   159: 126-35.

  • Jantsch J, Gerlach RG, Ensser A, Dahesh S, Popp I, Heeg C, Bleiziffer O, Merz T, Schulz T, Horch RE, Bogdan C, Nizet V, van der Linden M   (2013).   Severe soft tissue infection caused by a non-beta-hemolytic Streptococcus pyogenes strain harboring a premature stop mutation in the sagC gene.   J Clin Microbiol.   51: 1962-5.

  • Brulois K., Chang H., Lee A., Ensser, A., Wong LY., Toth Z., Lee S.H., Lee H.-R., Myoung J., Ganem D., Oh T.-K., Kim J.F., Gao S.J. and Jung J.U.   (2012).   Construction and manipulation of a new Kaposi's sarcoma-associated herpesvirus Bacterial artificial chromosome clone.   J Virol.   [in press] .

  • Hahn A., Kaufmann J., Wies E., Naschberger E., Ivlev J., Schmidt K., Holzer A., Schmidt M., Chen J., König S., Ensser, A., Myoung J, Brockmeyer N.H., Stürzl M., Fleckenstein B., Neipel F.   (2012).   The Ephrin Receptor Tyrosine Kinase A2 is a Cellular Receptor for Kaposi's Sarcoma-Associated Herpesvirus.   Nature Medicine 18: 961-966.

  • Lehner M., Kellert B., Proff J., Schmid M., Diessenbacher P., Ensser, A., Dörrie J., Schaft N., Leverkus M., Kämpgen E., Holter W.   (2012).   Autocrine TNF is critical for the survival of human dendritic cells by regulating BAK, BCL-2 and FLIPL.   J Immunol. 188: 4810-8.

  • Dey Mazumder E., Jardin C, Vogel B., Heck E, Scholz B., Lengenfelder D., Sticht H., Ensser, A.   (2012).   A molecular model for the differential activation of STAT3 and STAT6 by the herpesviral oncoprotein Tip.   PLoS ONE 7: e34306.

  • Lehner M., Götz G., Schaft N., Dörrie J., Full F., Ensser, A., Muller Y., Cerwenka A., Abken H., Parolini O., Ambros P, Kovar H., Holter W.   (2012).   NKG2D-Ligands for Redirecting T Cells to Ewing's Sarcoma Family of Tumors and Rapid Ligand-Induced Loss of RNA-Transferred Receptor.   PLoS ONE 7: e31210.

  • Full F.*, Reuter N.*, Scholz B., Stamminger T.*, Ensser, A.*   (2012).   Herpesvirus saimiri antagonizes nuclear domain 10-instituted intrinsic immunity via an ORF3-mediated selective degradation of the cellular protein Sp100.   *these authors contributed equally.   J Virol. 86: 3541-53.

  • Zielke K., Full F., Teufert N., Schmidt M., Müller-Fleckenstein I., Alberter A., Ensser, A.   (2012).   The insulator protein CTCF binds to the orf73/LANA promoter region of Herpesvirus saimiri and is involved in conferring episomal stability in latently infected human T cells.   J Virol. 86: 1862-73.

  • Kingston D., Chang H., Ensser, A., Lee H.R., Lee J., Lee S.H., Jung J.U., Cho N.H.   (2011).   Inhibition of retromer activity by herpesvirus saimiri tip leads to CD4 downregulation and efficient T cell transformation.   J Virol. 85: 10627-38.

  • Alberter B., Vogel B., Lengenfelder D., Full F., Ensser, A.   (2011).   Genome-wide histone acetylation profiling of Herpesvirus saimiri in human T cells upon induction with a histone deacetylase inhibitor.   J Virol. 85: 5456-64.

  • Toptan T, Ensser, A. and Fickenscher H.   (2010).   Rhadinovirus vector-derived human telomerase reverse transcriptase expression in primary T cells.   Gene Therapy 17: 653-61.

  • Full F., Lehner M., Thonn V., Götz G., Scholz B., Kaufmann K.B., Mach M., Abken H., Holter W. and Ensser, A.   (2010).   T cells engineered with a CMV specific chimeric immunoreceptor.   J Virol. 84: 4083-4088.

  • Vogel B., Full F., Alberter B., Linden C. and Ensser, A.   (2010).   Episomal replication timing of gamma-herpesviruses in latently infected cells.   Virology 400: 207-214.

  • Lehner, M., Grillhoesl, C., Full, F., Weller, P., Müller-Fleckenstein, I., Schmidt, M., Fleckenstein, B., Holter, W. and Ensser, A.   (2009).   Transformation efficiency by Herpesvirus saimiri is not a limiting factor in clonal CD8pos T cell outgrowth.   Virology 388, 15-20.

  • Alberter, B., and Ensser, A.   (2007).   Histone modification pattern of the T cellular Herpesvirus saimiri genome in latency.   J Virol 81, 2524-2530.

  • Heck, E., Friedrich, U., Gack, M. U., Lengenfelder, D., Schmidt, M., Müller-Fleckenstein, I., Fleckenstein, B., Ensser, A. and Biesinger, B.   (2006).   Growth transformation of human T cells by herpesvirus saimiri requires multiple Tip-Lck interaction motifs.   J Virol 80, 9934-9942.

  • Wieser, C., Stumpf, D., Grillhösl, C., Lengenfelder, D., Gay, S., Fleckenstein, B. and Ensser, A.   (2005).   Regulated and constitutive expression of anti-inflammatory cytokines by non-transforming Herpesvirus saimiri vectors.   Gene Therapy 12, 395-406.

  • Ensser, A., Glykofrydes, D., Niphuis, H., Kuhn, E.M., Rosenwirth, B., Heeney, J.L., Bruder, J., Niedobitek, G., Müller-Fleckenstein, I. and Fleckenstein, B.   (2001).   Independence of herpesvirus induced T-cell lymphoma from viral cyclinD homologue.   J Exp Med 193, 637-642.

 

Suggested reviews

  • Ensser, A. (2008). Simian Gammaherpesviruses. Encyclopedia of Virology, 3rd Ed., Academic Press/Elsevier 4, 589-594.

  • Fleckenstein, B. and Ensser A. (2007). Chapter 60: EBV and KSHV related viruses in non-human primates. Human Herpesviruses: Biology, Therapy and Immunoprophylaxis., 1076-1092.