ROLE OF EARLY AND LATE REPLICATION EVENTS IN INDUCTION OF APOPTOSIS BY BACULOVIRUSES

Autographa californica nuclear polyhedrosis virus (AcMNPV) mutants tha t lack the apoptotic suppressor gene p35 cause apoptosis in Spodoptera frugiperda SF21 cells. To identify a viral signal(s) that induces pro grammed cell death, we first defined the timing of apoptotic events du ring infection. Activation of a P35-inhibitable caspase, intracellular fragmentation of host and AcMNPV DNA, and cell membrane blebbing coin cided with the initiation of viral DNA synthesis between 9 and 12 h af ter infection and thus suggested that apoptotic signaling begins at or before this time. Virus entry was required since binding of budded vi rus to host cell receptors alone was insufficient to induce apoptosis. To therefore determine the contribution of early and late replication events to apoptotic signaling, we used the AcMNPV mutant ts8 with a t emperature-sensitive lesion in the putative helicase gene p143. At the nonpermissive temperature at which viral DNA. synthesis was condition ally blocked, ts8 caused extensive apoptosis of the SF21 cell line P35 (76D), which dominantly interferes with anti-apoptotic function of vir al P35. Confirming that apoptosis can be induced in the absence of nor mal viral DNA synthesis, parental SF21 cells also underwent apoptosis when infected with a ts8 p35 deletion mutant at the nonpermissive temp erature. However, maximum levels of ts8 p35 deletion mutant-induced ap optosis required a temperature-sensitive event(s) that included the in itiation of viral DNA synthesis. Collectively, these data suggested th at baculovirus-induced apoptosis can be triggered by distinct early (p re-DNA synthesis) and late replicative events, including viral DNA syn thesis or late gene expression.