Characterization of herpes simplex virus-containing organelles by subcellular fractionation: Role for organelle acidification in assembly of infectious particles
Ca. Harley et al., Characterization of herpes simplex virus-containing organelles by subcellular fractionation: Role for organelle acidification in assembly of infectious particles, J VIROLOGY, 75(3), 2001, pp. 1236-1251
The cytoplasmic compartments occupied by exocytosing herpes simplex virus (
HSV) are poorly defined. It is unclear which organelles contain the majorit
y of trafficking,virions and which are occupied by virions on a productive
rather than defective assembly pathway. These problems are compounded by th
e bet that HSV-infected cells produce virus continuously over many hours. A
ll stages in viral assembly and export therefore coexist, making it impossi
ble to determine the sequence of events and their kinetics. To address thes
e problems, we have established assays to monitor the presence of capsids a
nd em eloped virions in cell extracts and prepared HSV-containing organelle
s from normally infected cells and from cells undergoing a single synchroni
zed wave of viral egress. We find that, in both cases, HSV particles exit t
he nucleus and accumulate in organelles which cofractionate with the trans-
Golgi network (TGN) and endosomes. In addition to carrying enveloped infect
ious virions in their lumen, HSV-bearing organelles also displayed nonenvel
oped capsids attached to their cytoplasmic surface. Neutralization of organ
ellar pH by chloroquine or bafilomycin A resulted in the accumulation of no
ninfectious enveloped particles. We conclude that the organelles of the TGN
/endocytic network play a hey role in the assembly and trafficking of infec
tious HSV.