Cell entry by reoviruses requires a large, transcriptionally active subvirion particle to gain access to the cytoplasm. The features of this particle have been the subject of debate, but three primary candidates-the infectious subvirion particle (ISVP), ISVP*, and core particle forms-that differ in whether putative membrane penetration protein μ1 and adhesin σ1 remain particle bound have been identified. Experiments with antibody reagents in this study yielded new information about the steps in particle disassembly during cell entry. Monoclonal antibodies specific for the δ region of μ1 provided evidence for a conformational change in μ1 and for release of the δ proteolytic fragment from entering particles. Antiserum raised against cores provided evidence for entry-related changes in particle structure and identified entering particles that largely lack the δ fragment inside cells. Antibodies specific for σ1 showed that it is also largely shed from entering particles. Limited coimmunostaining with markers for late endosomes and lysosomes indicated the particles lacking δ and σ1 did not localize to those subcellular compartments, and other observations suggested that both the particles and free δ were released into the cytoplasm. Essentially equivalent findings were obtained with native ISVPs and highly infectious recoated particles containing wild-type proteins. Poorly infectious recoated particles containing a hyperstable mutant form of μ1, however, showed no evidence for the in vitro and intracellular changes in particle structure normally detected by antibodies, and these particles instead accumulated in late endosomes or lysosomes. Recoated particles with hyperstable μ1 were also ineffective at mediating erythrocyte lysis in vitro and promoting α-sarcin coentry and intoxication of cells in cultures. Based on these and other findings, we propose that ISVP* is a transient intermediate in cell entry which mediates membrane penetration and is then further uncoated in the cytoplasm to yield particles, resembling cores, that largely lack the δ fragment of μ1.
|Original language||English (US)|
|Number of pages||15|
|Journal||Journal of virology|
|State||Published - Dec 1 2003|
ASJC Scopus subject areas
- Insect Science