Edwards, C.S.; (2005) The roles of vaccinia virus proteins in kinesin-dependent microtubule transport. Doctoral thesis , University of London.

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Abstract

Vaccinia virus is a good model system for studying host cell biology and enhancing our understanding of microtubule-based transport. Whilst the key viral proteins involved in microtubule-based transport of intracellular enveloped virus (IEV) have been identified, their exact mechanistic relationship to each other and to microtubule motor proteins has not been fully established. This thesis aims to address how the viral protein F12L and kinesin-1 are recruited to IEV particles and how the viral proteins A33R and A36R are also involved in this process. It is known that the movement of IEV particles, from the perinuclear viral factories to the plasma membrane, is microtubule-dependent and involves the A36R protein as well as the recruitment of kinesin-l. At the plasma membrane, actin tail formation has been shown to be dependent on phosphorylation of A36R. However, A36R is dependent upon an interaction with the A33R protein in order to be recruited to the virion. It has also been shown that A33R is heavily phosphorylated at serine residues but its role is currently unknown. The initial part of my studies examined the potential role of phosphorylation of A33R and its effect on viral egress. No effect on viral transport was observed upon dephosphorylation of A33R. The viral protein F12L also has a proposed role in microtubule-dependent transport, as an accumulation of particles is observed in the region of the viral factories and transport to the cell surface does not occur in the absence of the protein. However, the mode of interaction of F12L with IEV and its association with kinesin-l is unclear. The precise structure of F12L is also unknown. In order to investigate the relationship between F12L, A36R, A33R and kinesin-l, a new range of tools was developed, including: a F12L antibody, GFP-F12L fusion constructs, deletion mutants to determine which regions of F12L are involved in virus transport, constructs to investigate whether F12L undergoes any post-translational modification event(s), as well as designing F12L and A33R recombinant viruses. No direct interaction has been established between F12L and A36R, or A33R, or kinesin-l.

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