G·A·B·B·A

Herpesvirus particles are complex structures that have four generic properties: a core enclosing a linear double stranded DNA virus genome, an icosahedral capsid, a complex tegument layer surrounding the capsid, and a lipid bilayer membrane that forms the virion envelope and contains numerous viral glycoproteins. Our knowledge of the assembly and egress of this complex structure is superficial.
According to the two-step envelopment pathway of virion maturation and egress, HSV-1 nucleocapsids assemble in the nuclei of infected cells and acquire an envelope by budding through the inner nuclear membrane. After released into the cytoplasm by deenvelopment at the outer nuclear membrane, secondary envelopment is thought to take place at the trans-Golgi network compartment, and finally virions are released into extracellular space by fusion with the plasma membrane. Although generally accepted, this pathway is not consensual.
This two-step envelopment pathway implies that the final envelope is gained from a subcellular compartment distinct from that of the nuclear membrane/ER. To address this question an immunogold electron microscopy approach was used to determine the tegument composition of primary enveloped virions, which lie between both leaflets of the nuclear membrane. VP13/14 and VP22 proteins were not detected in virus particles in the perinuclear space and were only present in mature extracellular virions, whilst VP16 protein was acquired prior to primary envelopment of the virus at the inner nuclear membrane. To gain further insight into the morphogenesis of HSV-1 virions, recombinant viruses lacking VP1/2 protein - an essential structural component of the virus tegument and conserved in all herpesviruses - and expressing GFP/YFP-tagged tegument proteins VP16, VP22 and VP13/14 were generated. Immunogold electron microscopy studies showed that the absence of VP1/2 did not prevent the incorporation of VP16GFP into primary enveloped virions, whereas neither VP22GFP nor VP13/14YFP were detected on capsids in the cytoplasm, suggesting that VP1/2 may play a role in the recruitment of these other tegument proteins into the assembling HSV-1 virus particle. The evidence presented in this thesis supports the two-step envelopment model of egress in which enveloped virus particles in the perinuclear space fuse with the outer nuclear membrane, release naked nucleocapsids into the cytoplasm, acquiring the final envelope from a post-ER compartment.