FIGURE 8.

Expression of flavivirus NS4A alone leads to protection similar to that provided by infection with live flavivirus. A, individual expression of any of the 10 Dengue-2 structural and nonstructural genes does not lead to significant death in MDCK renal epithelial cells. Dengue-2 NS4A is the only gene that protects against cell killing by CPT, similar to live virus infection. NS2A and NS5 each allow slightly more death by CPT than mock − infected cells. Individual expression of empty plasmid vector, the structural protein capsid or core (C), premembrane (prM), and envelope (E) proteins. NS1, NS2B, NS3, or NS4B has no effect on CPT-induced death. B, infection with live Modoc virus infection or individual expression of Modoc NS4A also protects cells against CPT-induced death. Expression of NS2A, NS4A, or NS4B does not lead to significant cell killing in MDCK cells (<10%). Modoc NS4A expression protects in a manner similar to live virus infection. Modoc NS2A expression allowed slightly more killing than in mock − infected cells. Expression of the empty plasmid vector or Modoc NS4B had no effect on CPT-induced killing. C, NS4A expression from either Dengue-2 or Modoc virus protects against a DNA-damaging agent (CPT) and a kinase inhibitor (STS). Up-regulation of autophagy by rapamycin treatment protected against death by CPT regardless of expression of NS4A from either virus. Inhibition of autophagy by 3MA treatment abolishes Dengue-2 and Modoc NS4A-induced protection. Inhibition of autophagy by wortmannin also eliminates protection against STS-induced cell death induced by NS4A from either virus.