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Title: Biological properties of the X-gene product of hepatitis B virus
Author: Rossner, Michael T.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1991
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Several biological properties of the X-gene product of hepatitis B virus (HBV) were investigated to begin to elucidate its function in the viral life-cycle and its contribution to the physiological consequences of HBV infection. One of these consequences is a humoral immune response mounted by the host that is directed against the X-gene product (HBxAg). The antigenicity of segments of HBxAg was investigated by expressing them as fusion proteins in E.coli. Cross-reactivity of these products was determined with antisera from an HBV infected chimpanzee and from a rabbit inoculoated with HBxAg produced in E.coli. These polyclonal antisera contained antibodies to all segments of HBxAg, although much weaker reactivity with a fusion protein containing only the middle third of the antigen was observed with both antisera. A striking serological feature of HBV infection is the vast amount of viral surface antigen (HBsAg) secreted from infected cells, and the contribution of HBxAg to this high level of HBsAg expression was investigated. Human hepatoma cells transfected with a plasmid construction containing the transcription units for the preS2 and surface (preS2/S) mRNAs and the X mRNA secreted HBsAg into the culture medium. A frameshift mutation in the X gene greatly reduced the production of HBsAg. The mutation could be complemented by contransfection with a plasmid containing the X structural gene under control of the SV40 early promoter. Levels of HBsAg production were directly related to the amount of preS2/S mRNA produced showing that HBxAg can modulate expression of this gene. HBsAg polypeptides containing preS domains are secreted by HBV infected hepatocytes at a low level relative to the major-S polypeptides which contain only the surface domain. HBsAg did not influence the relative levels of the various forms of HBsAg polypeptide produced by hepatoma cells transfected with a plasmid encoding both preS domains. HBsAg production was reduced upon cotransfection of cells with a plasmid construction containing the HBV enhancer indicating that expression of HBsAg is influenced by proteins binding to this region. However, HBxAg could still exert its transactivating effect in the presence of the competitor plasmid, indicating that this effect can be mediated through DNA sequences outwith the enhancer region. The ability of HBxAg to function as a protein serine/threonine kinase was investigated to determine whether this polypeptide may be the source of the endogenous kinase activity associated with HBV particles. In addition, this enzymatic function could account for the ability of HBxAg to modulate transcription. No protein kinase activity was detected for HBxAg expressed in E.coli as a fusion protein with HBcAg or for HBxAg expressed in hepatoma cells.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available