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Immature particles in the perinuclear space are then proposed to bud through the
outer leaflet of the nuclear membrane, loosing their translocation membrane and
becoming nonenveloped cytoplasmic particles (Fig. 7, step 2). Although this gen-
eral pathway appears to be shared by all herpes viruses, there is less consensus
about where the tegument proteins are added.
With regard to the predominant tegument proteins of CMV, three sets of
observations are compatible with their proposed addition outside the nucleus
(Fig. 7, step 3). First, electron microscopy shows that cytoplasmic capsids have a
thick fibrillar coating (deduced to be tegument proteins) that is entirely absent from
nuclear capsids (Figs. 1C, 1D, 6; Fig. 1 in Gibson 1993). Second, SDS-PAGE
analyses show that capsids recovered from the cytoplasm of infected cells contain
the predominant tegument proteins, whereas those from the nucleus do not (Gibson
1981). And third, immunofluorescence studies show accumulations of three abundant
tegument proteins (pUL83, pUL32, pUL99) in assembly compartments juxtaposed
to the nucleus, but outside of it (Scholl et al. 1988; Sanchez et al. 2000a, 2000b)
(reviewed in Eickmann et al. 2006). None of these observations, however, rule out the
possibility that some or all of the same tegument proteins bind to capsids within
the nucleus or perinuclear space (Hensel et al. 1995; Nii et al. 1998) and are rapidly
translocated with the capsid into the cytoplasm where they accumulate to levels
more readily detected. Compatible with this possibility, recent studies show that
specific mutations in the tegument proteins pUL36 (VP1/2) of HSV, or pUL32
(basic phosphoprotein/pp150) of HCMV, result in their accumulation within the
nucleus (O'Hare and Abaitua 2006; J. Wang and W. Gibson, unpublished data from
studies using mutant viruses encoding CysCysProGlyCysCys-tagged pUL32
detected in live, infected cells with the biarsenical dye FIAsH).
Once in the cytoplasm and completely tegumented, the capsids bud into cyto-
plasmic vesicles or tubules to acquire their final envelope (e.g., Fig. 7, step 4). This
process required pUL99; in its absence, tegumented capsids accumulate in the
cytoplasm (Silva et al., 2003). Several changes appear to accompany this process;
including compression or tightening of the tegument and thickening of the mem-
brane with apparent elaboration on its luminal surface. These changes occur where
the membrane and capsid are in close proximity (e.g., Fig. 1d), suggesting confor-
mational or compositional changes in both layers as envelopment proceeds. Late
maturational events such as phosphorylation by the virion-associated kinase(s)
(Roby and Gibson 1986; Nogalski et al. 2007), carbohydrate processing, and pos-
sible redistribution of envelope and tegument constituents, are likely to occur as the
particle completes its egress and is modified to increase its efficiency as an entry
vessel for delivering the viral DNA to the next cell.
The recently discovered ubiquitin-specific cysteine protease (DUB) activity,
present and functional in virions at the amino end of the high-molecular-weight
tegument protein pUL48 of HCMV and pUL36 of HSV (Kattenhorn et al. 2005;
Schlieker et al. 2005; Wang et al. 2006), may be involved in these late events, or
have a role at the outset of infection, or both. Viruses mutated in the catalytically
critical Cys41 or His163 residues of the HCMV DUB replicate at a reduced efficiency
relative to wild type virus, indicating an important even if not absolutely essential
role for this new virion-associated enzyme (Wang et al. 2006).
