Rhadinovirus Rezeptoren - Struktur, Signaltransduktion und Zelltropismus
Zusammenfassung der Projektergebnisse
The Kaposi’s sarcoma-associated herpesvirus is associated with a significant disease burden in Sub-Sahara Africa and in immunocompromised individuals world-wide. We characterized the interaction of the Eph family of receptor tyrosine kinases with the gH/gL glycoprotein complex of Kaposi’s sarcoma-associated herpesvirus (KSHV) and the related rhesus monkey rhadinovirus (RRV), an animal model virus for KSHV. The gH/gL glycoprotein complex resides in the virion membrane, binds cellular receptors, and controls the fusion process of the viral and the host membrane. We further identified another family of receptors for RRV gH/gL, the Plexin domain-containing proteins 1 and 2 (Plxdc1/2), which are exclusively used by RRV but not by the human KSHV. Using mutagenesis and structural modelling we then identified defined regions on gH/gL that mediate binding to both the Eph family receptors and to Plxdc1/2. We used this information to create virus mutants that cannot use these receptors. Analysis of these mutant viruses demonstrated that Ephs function at an post-attachment step, i.e. after the virus has bound to the target cells, and that Eph-mediated infection is important for KSHV infection of all cell types tested, and not only for infection by cell-free virus but also for cell-to-cell transmission. We found that the spectrum of Ephs that is used by KSHV in this setting also encompasses EphA5 and most prominently EphA7, and that Eph-mediated cell-to-cell transmission is highly efficient. These findings and results by other groups also suggested that the KSHV gH/gL complex is an excellent antigen to elicit neutralizing antibodies. Indeed, we were able to elicit potently neutralizing antibody responses by immunizing mice with different preparations of the gH/gL complex as recombinant protein. Our results indicated neutralization through inhibition of receptor binding and through additional mechanisms and may lay the groundwork for the rational design of a KSHV vaccine. Our discovery of the Plxdc1/2 as alternative gH/gL receptors for RRV is compatible with findings that we made in the rhesus macaque animal model, where an RRV mutant that cannot infect via Eph receptors was still able to establish persistent infection, likely using Plxdc1/2 to gain access to the B cell compartment in vivo. This was fundamentally different from our findings in most B cell lines that were tested in vitro and that were infected in an almost exclusively Eph-dependent manner. These findings highlight that cell culture models may only have limited predictive value and that ultimately tissue tropism of viruses should be studied in animal models. We also found that such a receptor-restricted RRV still elicits potent cellular immune responses when used as a vaccine vector to present antigens derived from the simian immunodeficiency virus, an animal model virus for the human immunodeficiency virus (HIV). This is potentially important for the development of a live-attenuated KSHV vaccine or RRV/KSHV-based vaccine vectors against HIV/AIDS. Analyzing virus entry and cell-cell fusion mediated by KSHV and RRV glycoproteins, we found that KSHV and RRV are restricted by interferon-induced transmembrane proteins (IFITMs) in a cell-specific manner. IFITMs can interfere with membrane fusion, and we found that KSHV inhibition by IFITMs was more pronounced in a cell line where KSHV particles colocalized with IFITMs at early timepoints of infection than in endothelial cells where there was no early colocalization. KSHV may therefore escape IFITM-mediated restriction in endothelial cells by avoiding spatial proximity to these proteins during entry. We further confirmed previous findings on pH dependence of KSHV entry and found that membrane fusion by KSHV glycoproteins is highly dependent on vesicle pH. Finally, we made use of a genome-wide, lentiviral CRISPR library and next-generation sequencing and identified genes whose knockout renders cells resistant to RRV infection. This approach yielded two novel, so-far uncharacterized host factors that are conserved between KSHV and RRV and that mediate infection by both viruses.
Projektbezogene Publikationen (Auswahl)
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EphA7 functions as receptor on BJAB cells for cell-to-cell transmission of the Kaposi's sarcoma-associated herpesvirus (KSHV) and for cell-free infection by the related rhesus monkey rhadinovirus (RRV). Journal of Virology. 2019 May 22
Großkopf AK, Schlagowski S, Hörnich BF, Fricke T, Desrosiers RC, Hahn AS
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A Recombinant Rhesus Monkey Deleted of Glycoprotein L Establishes Persistent Infection of Rhesus Macaques and Elicits Conventional T Cell Responses. Journal of Virology. 2020 Jan 6;94(2)
Hahn AS, Bischof GF, Großkopf AK, Shin YC, Domingues A, Gonzalez-Nieto L, Rakasz EG, Watkins DI, Ensser A, Martins MA, Desrosiers RC
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Interferon-Induced Transmembrane Proteins Inhibit Infection by the Kaposi’s Sarcoma- Associated Herpesvirus and the Related Rhesus Monkey Rhadinovirus in a Cell-Specific Manner. mBio. 2021 Dec 21;12(6):e0211321
Hörnich BF, Großkopf AK, Dcosta CJ, Schlagowski S, Hahn AS
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Plxdc family members are novel receptors for the rhesus monkey rhadinovirus (RRV). PLoS Pathogens. 2021 Mar 03; 17(3): e1008979
Großkopf AK, Schlagowski SC, Ensser A, Desrosiers RC, Hahn AS
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SARS-CoV-2 and SARS-CoV spikemediated cell-cell fusion differ in the requirements for receptor expression and proteolytic activation. Journal of Virology. 2021 Feb 19;JVI.00002-21
Hörnich BF, Großkopf AK, Schlagowski S, Tenbusch M, Kleine-Weber H, Neipel F, Stahl-Hennig C, Hahn AS
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Antibodies Targeting KSHV gH/gL Reveal Distinct Neutralization Mechanisms. Viruses, 2022, 14 (3), 541
Fricke T, Großkopf AK, Ensser A, Backovic M, Hahn AS
