Intracellular disassembly of infectious rotavirus particles by depletion of Ca super(2) super(+) sequestered in the endoplasmic reticulum at the end of virus cycle

Rotavirus infection is characterized by a number of Ca super(2) super(+) dependent virus-cell interactions. The structure of rotavirus triple-layered particles (TLP) is dependent on Ca super(2) super(+) concentration. Acquisition of the capsid outer layer requires a high Ca super(2) super(+) concentration inside the ER. Infection modifies Ca super(2) super(+) homeostasis of the cell, increasing ER Ca super(2) super(+) content, which may be advantageous to virus replication. We studied the role of sequestered Ca super(2) super(+) on the stabilization of already mature viral particles within the ER. Thapsigargin (TG), a SERCA pump inhibitor, added for 30min at the end of infection depleted ER Ca super(2) super(+) and reduced the titer of already mature TLP accumulated in the cell. Another inhibitor, cyclopiazonic acid, and two Ca super(2) super(+) ionophores (A23187 and ionomycin) in the presence of EGTA had similar effects. TG eliminated the peak of radiolabeled TLP, increasing that of DLP in CsCl gradients. Electron microscopy revealed accumulation of clustered particles in the ER, which had lost their integrity. The [Ca super(2) super(+)] in the ER of infected cells is important for virus maturation and for maintaining the integrity of mature TLP. Viral particles in this compartment may be potentially infectious, already containing VP7 and VP4.