Reovirus Activates a Caspase-Independent Cell Death Pathway

Berger, Angela K.; Danthi, Pranav

Reovirus induces RIP1-dependent necrosis. One such caspase-independent cell death pathway requires the kinase activity of RIP1 (39, 40) . To assess whether the caspase-independent cell death pathway activated by reovirus is dependent on the kinase activity of RIP1, we determined the capacity of reovirus to induce cell death following treatment of cells with necrostatin-1 (Nec-1), an inhibitor of RIP1 (41) . We found that reovirusinduced cell death was blocked by Nec-1 (Fig. 4A ). Simultaneous treatment of cells with Nec-1 and a pan-caspase inhibitor did not further diminish cell death, suggesting that a RIP1 kinasedependent form of cell death is the primary mechanism of cell death in these cells (Fig. 4A ). The effect of Nec-1 on reovirusinduced cell death was independent of its effect on the blockade of indoleamine 2,3-dioxygenase (IDO), an additional target of Nec-1 (42, 43) , since direct inhibition of IDO using 1-methyl-Ltryptophan did not prevent reovirus-induced cell death (see Fig. S2A in the supplemental material). The capacity of reovirus to establish infection and grow in Nec-1-treated cells was not affected ( Fig. S2B and C). Nec-1 also did not diminish caspase-3/7 activity in T3D-infected cells (Fig. S2D) . These data suggest that RIP1 kinase activity is required for reovirus-induced cell death.