The role of RL13 in human cytomegalovirus pathogenesis

Human cytomegalovirus (HCMV) is the leading infectious cause of congenital malformation. In vitro propagation of HCMV strains that authentically represent clinical virus are required to facilitate effective research. However, this is problematic, since efficient replication of clinical HCMV strains in vitro is achieved only after mutations have been selected. The first gene to mutate is consistently RL13. RL13 encodes a virion envelope protein that suppresses release of infectious virus from cells. Despite mutation in cell culture it is conserved in clinical strains, indicating that it is important in vivo. However, it is unclear how RL13 inhibits virus in vitro, or what advantage it provides in vivo. RL13 expression did not affect genome replication or virion protein production, however it reduced the infectivity of released virions. Mass spectrometry revealed that RL13 expression altered the composition of released virions, however there was no obvious reason for these changes to effect infectivity. During virus spread, RL13 promoted cell-cell transfer of virus, reducing the ability of neutralising antibodies to inhibit virus spread in fibroblasts, however this effect was masked by UL128L expression in other cell types. Genetic analysis of RL13 from clinical strains revealed that RL13 sequences clustered into 9 distinct clades, with sequences from different clades retaining the ability to inhibit the release of infectious virus. A RL13 homologue from Rhesus CMV also restricted the release of infectious virus. This conservation of function suggests that the phenotype is not an artefact of in vitro culture conditions. RL13 on the cell surface bound human IgG, however it did not inhibit antibody-dependent cellular cytotoxicity during infection. This thesis has delivered insights into the role of RL13 during infection, has demonstrated the importance of using more clinically relevant RL13+ HCMV strains and will aid future research using such HCMV strains