Studies on MHC class I-mediated entry of equine herpesvirus-1 into cells

Equine herpesvirus 1 (EHV-1) is an endemic virus that affects horse populations worldwide, causing respiratory disease, abortion and a serious neurologic disease known as encephalomyelitis. Outbreaks of EHV-1 encephalomyelitis can have a severe economic impact on farms, racetracks and veterinary hospitals, and currently available vaccines are not effective to protect against EHV-1 encephalomyelitis. Histological analyses of affected horses indicate that viral infection of vascular endothelial cells leads to damage of the central nervous system due to the resulting shortage of blood supply. Although infection of these cells is closely associated with the development of encephalomyelitis, the molecular mechanisms underlying this association remain poorly understood.Alphaherpesviruses enter target cells through sequential multistep processes. Following the initial attachment of the viruses to the cell surface, binding of viral glycoproteins to cell surface receptors triggers fusion of the viral envelope with the cell membrane, resulting in the release of viral capsid (containing viral genome) into the cytoplasm. EHV-1 attaches to cell surface using an interaction between viral glycoprotein C (gC) and cell surface heparan sulfate. Although the role of gC is important for effective infection, it does not trigger viral entry into cells. Glycoprotein D (gD) of EHV-1 is known to be essential for EHV-1 entry into cells. However, functional gD receptor that mediates EHV-1 entry into equine central nervous system (CNS) endothelial cells remains unidentified.In chapter 1, I performed functional cloning using an equine brain microvascular endothelial cell cDNA library and identified that equine major histocompatibility complex (MHC) class I heavy chain conferred susceptibility to EHV-1 infection in mouse NIH3T3 cells, which are resistant to EHV-1 infection. I investigated the role of equine MHC class I in EHV-1 infection and revealed that equine MHC class I acts as an entry receptor for EHV-1 through its binding to EHV-1 gD. I further investigated the role of MHC class I in EHV-1 entry into different types of equine cells.In chapter 2, I constructed equine MHC class I heavy chain specific-RNA probe and performed Northern hybridization to examine the level of equine MHC class I gene expression in the major organs of the adult horse body. The localization of equine MHC class I mRNA in brain tissue was investigated by in situ hybridization.In this thesis, I demonstrated equine MHC class I is a bona fide receptor of EHV-1 entry into equine cells. This study provides new insights into the mechanism of EHV-1 entry into host cells and a potential way to treat and prevent this infectious disease.