Biology Reference
In-Depth Information
the E330K VLP from undergoing the calcium-dependent structural
shift necessary for binding to the cell receptor, the major histocom-
patibility complex (MHC) class I molecule, leading to the adsorption
defect. Analyzing the structural difference between the mutant E330K
VLP and the wild-type virion could help delineate the virus site or
epitope responsible for attachment to MHC class I. Another point
mutant E157K, the site-2 calcium mutant, about 10% larger in diam-
eter than the wild type, was able to enter cells, but this did not lead
to T-antigen expression.
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Cell-internalized E157K DNA effectively
coimmunoprecipitated with anti-VP1 antibody, but little of the DNA
did so with anti-VP3 antibody, and none was detected in anti-importins
immunoprecipitate. Yet a substantial amount of VP3 was present in
anti-VP1 immune complexes, suggesting that internalized E157K
particles are ineffective at exposing VP3. These results show that
mutant E157K infection is blocked at a stage prior to the interaction
of VP3 nuclear localization signal with importins, consistent with a
role for calcium-binding site-2 in post-entry steps leading to the nuclear
import of the infecting SV40.
Virion Disassembly and Viral Genome
Entrance into the Nucleus
SV40 is unusual among animal viruses in that it enters cells through
caveolae, and the internalized virus accumulates in a smooth endo-
plasmic reticulum (ER) compartment (Figs. 4 and 5). After associating
with caveolae, SV40 leaves the plasma membrane in small, caveolin-
1-containing vesicles.
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It then enters larger, peripheral organelles
with a non-acidic pH. Although rich in caveolin-1, these organelles
do not contain markers for endosomes, lysosomes, ER or Golgi, nor
do they acquire ligands of clathrin-coated vesicle endocytosis. After
several hours in these organelles, SV40 is sorted into tubular, cave-
olin-free membrane vesicles that move rapidly along microtubules,
and is deposited in perinuclear, syntaxin 17-positive, smooth ER
organelles. There exists a two-step transport pathway from the plasma-
membrane caveolar, through an intermediate organelle termed the
caveosome, to the ER. This pathway bypasses endosomes and the
