G. Griffiths et al., Structure and assembly of intracellular mature vaccinia virus: Isolated-particle analysis, J VIROLOGY, 75(22), 2001, pp. 11034-11055
In a series of papers, we have provided evidence that during its assembly v
accinia virus is enveloped by a membrane cisterna that originates from a sp
ecialized, virally modified, smooth-membraned domain of the endoplasmic ret
iculum (ER). Recently, however, Hollinshead et al. (M. Hollinshead, A. Vand
erplasschen, G. I. Smith, and D. J. Vaux, J. Virol. 73:1503-1517, 1999) arg
ued against this hypothesis, based on their interpretations of thin-section
ed material. The present article is the first in a series of papers that de
scribe a comprehensive electron microscopy (EM) analysis of the vaccinia In
tracellular Mature Virus (JAM and the process of its assembly in HeLa cells
. In this first study, we analyzed the IMV by on-grid staining, cryoscannin
g EM (SEM), and cryo-transmission EM. We focused on the structure of the IM
V particle, both after isolation and in the context of viral entry. For the
latter, we used high-resolution cryo-SEM combined with cryofixation, as we
ll as a novel approach we developed for investigating vaccinia IMV bound to
plasma membrane fragments adsorbed onto EM grids. Our analysis revealed th
at the IMV is made up of interconnected cisternal and tubular domains that
fold upon themselves via a complex topology that includes an S-shaped fold.
The vital tubules appear to be eviscerated from the particle during viral
infection. Since the structure of the MINI is the result of a complex assem
bly process, we also provide a working model to explain how a specialized s
mooth-ER domain can be modulated to form the IMV. We also present theoretic
al arguments for why it is highly unlikely that the IMV is surrounded by on
ly a single membrane.