Arriving at an in-depth understanding of equine herpesvirus type 1 and 4 (EHV-1 & EHV-4) disease: The role of mononuclear cells in viral pathogenesis

• Applicant: Professor Dr. Nikolaus Osterrieder, since 4/2020
• Subject Area: Veterinary Medical Science ; Virology
• Term: from 2013 to 2016
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 251507118

Equine herpesvirus type 1 (EHV-1) and EHV-4 are members of the Herpesviridae family, more specifically the Alphaherpesvirinae subfamily and the genus Varicellovirus. Both viruses are endemic in horse populations and can cause significant suffering of horses and economic losses to the equine industry following clinical disease that includes respiratory tract problems, abortions, and neuronal disorders. We took advantage of the close genetic relatedness between both viruses and exchanged essential genes between EHV-1 and EHV-4, which allowed us to evaluate viral genes in a way impossible for their relatives in the Alphaherpesvirinae subfamily. We were able to show that both EHV-1 and EHV-4 use the same entry receptor (MHC-I) to infect equine cells and that the glycoprotein (g)D ectodomain is responsible for MHC-I interaction but also defines tropism by interaction with non-MHC-I receptors on cells from non-equine origin. We showed also that EHV-1 and EHV-4 employ different entry pathways during infection of epithelial cells and interaction of viral gH with integrins results in a dramatic increase of cytosolic Ca2+ that precedes phosphatidylserine scrambling. Changes in lipid composition then facilitates virus fusion at the plasma membrane. Finally, only EHV-1-infected (but not EHV-4-infected) PBMC were efficiently tethered to and rolled over EC, a process that resulted in virus transfer to EC with gB and the US3 protein are key players to help EHV-1 cell-to-cell spread. It is our long-term goal to understand the entry mechanism of EHV-1 and EHV-4 as well as virus spread within the host, with an emphasis on the transfer of virus from PBMC to endothelial cells. We hypothesize that the higher pathogenic potential and efficient cell-to-cell spread of EHV-1 when compared to EHV-4 are determined by binding to different cellular receptors, different routes of entry, and hijacking of various cellular processes by modulating signaling. We further surmise that gD, gH/gL, gB, but also the US3 protein kinase play important roles in establishing productive infection by activating cellular molecules, including phospholipids and signaling proteins, next to engaging cell surface receptors.We will test our hypothesis in three specific aims:1- To determine the crystal structure of gD and define receptor binding domains 2- To investigate cellular and viral factors affecting virus entry 3- To investigate the mechanism of virus transmission from infected PBMC to EC

• DFG Programme: Research Grants
• Ehemaliger Antragsteller: Walid Azab, Ph.D., until 4/2020