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Title: Towards a pharmacology intervention for malignant catarrhal fever in cattle
Author: Mohammed Alethari, Ishtar Adana
ISNI:       0000 0004 7656 9906
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2019
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Malignant catarrhal fever (MCF) is a frequently lethal disease, which is characterised by high fever, depression, profuse nasal discharge, corneal opacity, as well as hyperaemic to ulcerative lesions in the mucosa of the respiratory tract. This leads to ocular and nasal discharge, diarrhoea. Severe inflammation of the conjunctival, oral, and nasal mucosa is accompanied by necrosis in the oral and nasal cavities sometimes extending into the oesophagus and trachea. MCF in general is sporadic, affecting individual animals within a group, but occasionally can cause high losses in the herd. MCF in domestic cattle is caused by two γ-herpesviruses namely ovine herpesvirus 2 (OvHV-2) and alcelaphine herpesvirus 1 (AlHV-1). AlHV-1 is endemic in wildebeest (Connochaetes taurinus) in Africa where it causes wildebeest associated malignant catarrhal fever (WA-MCF). Whereas OvHV-2 is endemic in sheep populations worldwide and causes sheep associated malignant catarrhal fever (SA-MCF). MCF is characterized by marked T cell hyperplasia and proliferation of unrestricted cytotoxic large granular lymphocytes (LGLs) which leads to necrosis of infiltrated tissues. These LGLs can be grown out in culture and contain OvHV-2 in a latent form. Little is known about the underlying molecular basis of MCF pathogenesis or what controls the differences in clinical outcome of infection in two closely-related host species. The overall aim of this project was to understand better the interactions between OvHV-2 and cellular proteins during latency to identify possible druggable targets that could be used for MCF therapy. All γ-herpesviruses express a protein during latency called latency-associated nuclear antigen (LANA). This protein in other herpesviruses is a master-regulator of virus latency. It tethers virus genome to host chromatin and ensures genome replication during latency as well as interacting with cellular proteins to control cell replication and also innate defence mechanisms. OvHV-2 has been shown to encode and express a LANA protein (oLANA) during latency in LGLs. The specific aim of this project was to determine the interactions of oLANA with cellular proteins which would give insight into pathogenesis as well as targets for therapeutic intervention. To do this, co-immunoprecipitatioin in HEK 293T cells was combined with label-free quantitative mass spectrometry to identify binding partners. A GFP-tagged recombinant oLANA lacking the central repetitive domain (GFP-oLANAΔ) was used as bait with GFP-trap technology to pull down cellular partners and then co-precipitants were analysed by mass spectrometry. Eight cellular proteins were identified as potential binding partners in this process. Of these, only histone H1 was validated by co immunoprecipitation and western blotting. While two others CCDC12 and Rad50 were shown to co-localise with oLANA by immunofluorescence and confocal microscopy. These results confirm, as other LANAs, a chromatin binding role for oLANA. They also give potential cellular targets for future investigation and therapeutic intervention.
Supervisor: Stewart, James Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral