ULTRASTRUCTURE OF KUNJIN VIRUS-INFECTED CELLS - COLOCALIZATION OF NS1AND NS3 WITH DOUBLE-STRANDED-RNA, AND OF NS2B WITH NS3, IN VIRUS-INDUCED MEMBRANE STRUCTURES
Eg. Westaway et al., ULTRASTRUCTURE OF KUNJIN VIRUS-INFECTED CELLS - COLOCALIZATION OF NS1AND NS3 WITH DOUBLE-STRANDED-RNA, AND OF NS2B WITH NS3, IN VIRUS-INDUCED MEMBRANE STRUCTURES, Journal of virology, 71(9), 1997, pp. 6650-6661
The subcellular location of the nonstructural proteins NS1, NS2B, and
NS3 in Vero cells infected with the flavivirus Kunjin was investigated
using indirect immunofluorescence and cryoimmunoelectron microscopy w
ith monospecific antibodies, Comparisons were also made by dual immuno
labelling using antibodies to double-stranded RNA (dsRNA), the putativ
e template in the flavivirus replication complex, At 8 h postinfection
, the immunofluorescent patterns showed NS1, NS2B, NS3, and dsRNA loca
ted in a perinuclear rim with extensions into the peripheral cytoplasm
, By 16 h, at the end of the latent period, all patterns had changed t
o some discrete perinuclear foci associated with a thick cytoplasmic r
eticulum, By 24 h, this localization in perinuclear foci was more appa
rent and some foci were dual labelled with antibodies to dsRNA, In imm
uno-gold-labelled cryosections of infected cells at 24 h, all antibodi
es were associated with clusters of induced membrane structures in the
perinuclear region, Two important and novel observations were made, F
irst, one set of induced membranes comprised vesicle packets of smooth
membranes dual labelled with anti-dsRNA and anti-NS1 or anti-NS3 anti
bodies, Second, adjacent masses of paracrystalline arrays or of convol
uted smooth membranes, which appeared to be structurally related, were
strongly labelled only with anti-NS2B and anti-NS3 antibodies, Paired
membranes similar in appearance to the rough endoplasmic reticulum we
re also labelled, but less strongly, with antibodies to the three nons
tructural proteins, Other paired membranes adjacent to the structures
discussed above enclosed accumulated virus particles but were not labe
lled with any of the four antibodies, The collection of induced membra
nes may represent virus factories in which translation, RNA synthesis,
and virus assembly occur.