Professor of Microbiology
Department of Microbiology
Perelman School of Medicine
University of Pennsylvania
203 A Johnson Pavilion
3610 Hamilton Walk
Philadelphia, PA 19104-6076
My lab focuses on the study of pathogenesis of coronaviruses. We have been studying the murine coronavirus, mouse hepatitis virus (MHV) for many years. MHV infection of mice provides a model system for the study of: 1) acute viral infection of the central nervous system (CNS) and both innate and acquired immune response to this infection; 2) a chronic persistent infection which proceeds in the presence of virus specific CD8T cells and provides one of the best models for human demyelinating diseases such as Multiple Sclerosis; 3) virus-induced hepatitis and 4) SARS like respiratory disease. Our long term goal is to elucidate the viral and cellular determinants of tropism and pathogenesis in the brain, the liver and the lung. In order to conduct these investigations, we have the important tools of a well-developed animal model system and two reverse genetic systems with which to manipulate the viral genome.
Currently we are carrying out the following studies:
1. Interaction of MHV with the type I interferon response. Interferon (IFN)-a/b is a very early host response, vital for the protection of the mouse from MHV infection in vivo. While MHV fails to induce type I IFN in tissue culture cell lines and some types of primary cells, we have found that MHV induces IFN-a/b in macrophage lineage cells and that induction requires expression of MDA5. We have also found that the basal level of expression of interferon stimulated genes differs in the CNS and the liver, the two main targets of MHV infection, with the liver seemingly more prepared for viral invasion, consistent with exposure of the liver to the environment. We found that the MHV ns2 gene encodes a phosphodiesterase activity and furthermore it is a liver specific virulence factor and an IFN antagonist that appears to specifically inhibit the OAS-RNase L pathway, a potent interferon stimulated antiviral pathway. RNase L is an important factor in the protection of mice from MHV induced hepatitis. Interestingly RNaseL does not seem to be an important factor in restriction of virus replication during acute infection of the CNS but may play a role in defense against the development of demyelination. Current studies are directed at further understanding the organ specific differences in host responses to MHV.
2. Role of receptor in MHV pathogenesis. The only known receptor for MHV is the carcinoembryonic antigen protein, CEACAM 1. However, the highly neurovirulent MHV-JHM strain spreads cell to cell in the absence of the expression of this receptor. Using transgenic CEACAM 1 deficient mice (ceacam 1a -/-), we have found that JHM can replicate and cause CNS disease and mortality in the absence of viral receptor. We are using a selection of MHV mutants and tropism variants in combination with primary cells derived from wild type and ceacam 1a-/- mice to elucidate the mechanism of viral neuron to neuron spread and the role of receptor independent spread in disease. We also plan to discover alternate receptors used by MHV in the absence of ceacam1a.
3. Virus induced demyelination. Following acute infection, MHV RNA genome persists in the CNS and expresses mRNA, in the absence of infectious virus; during this stage of infection, demyelination develops. Virus specific CD8T cells also persist during this late disease stage. We are investigating several aspects of persistent infection and the mechanisms leading to demyelinating disease. These include: the state of the viral genome and the integrity of viral mRNAs during persistent disease; the phenotype and functionality of the persisting CD8T cells and the possible role of RNaseL signaling in limiting demyelination.
4. MHV-1 and SARS coronavirus (SARS-CoV) induced severe respiratory disease. We are using our experience and knowledge about murine coronavirus to study the SARS associated virus. We are currently working with the MHV-1 model of SARS, in which a murine coronavirus produces similar pathology to SARS in a mouse model. One aspect of SARS induced pulmonary disease is downregulation of expression of the SARS receptor, angiotensin converting enzyme (ACE)-2, which in turns causes dysregulation of the rennin-angiotensin system which results in increased inflammation. We are expressing the SARS spike protein receptor binding domain from within the MHV-1 genome in order to study the contributions to pneumovirulence of dysregulation of the rennin angiotensin system, in the absence of SARS virus entry. We will then use the model to determine the efficacy of pharmacologic manipulation of the rennin-angiotensin system, by the use of ACE-1 inhibitors.
1. Organ specific interferon response to MHV. Investigation of the mechanism by which MHV ns2 inhibits the oligoadenylate synthetase (OAS)-ribonuclease (RNase) L interferon stimulated anti-viral pathway. This includes understanding the importance/role of RNaseL protection from hepatitis and demyelinating disease. This project involves studies of virus in primary cell types as well as in vivo experiments in mice. This project involves carrying out a wide variety of techniques, including viral pathogenesis, cell culture, cloning, in vitro protein expression and real time qPCR.
2. Mechanisms of MHV neurovirulence. Investigation of the special mechanism by which MHV spreads among neurons, independent of MHV receptor CEACAM 1a in vitro and in vivo during infection of the CNS. This project involves infections in primary neuronal cell cultures and in vivo with JHM and several tropism mutants and variants. We believe that the neuronal tropism of JHM is related to its inability to induce an effective CD8T cell response and this may be explored as well. Techniques include construction of recombinant viruses by reverse genetics techniques, infections in vivo and in in vitro neuronal cell culture as well as working with CD8T cells.
Ruth Elliott - Research Specialist
Judy Phillips- Postdoctoral Researcher
Ashley Wu- Undergraduate, Vagelos Program
Zachary Zalinger, Graduate Student
Ling Zhao- Postdoctoral Researcher
Rong Zhang, Visiting Graduate Student