ABSTRACTHerpes simplex virus 1 mutants lacking the gene encoding glycoprotein D (gD) and the gD normally present in the envelope of the virus (gD−/−stocks) or mutants lacking the gD gene but containingtrans-induced gD in their envelopes (gD−/+) cause apoptosis in human SK-N-SH cells. The gD−/−virions are taken up by endocytosis and are degraded, whereas gD−/+viruses replicate but produce gD−/−virus. Apoptosis is blocked by delivery of the gD gene intrans. Studies designed to test several hypotheses concerning the role of gD in apoptosis revealed the following. (i) gD−/−and gD−/+stocks induce fragmentation of cellular DNA in SK-N-SH, HEp-2, HeLa, and Vero cell lines. (ii) Chloroquine blocks apoptosis induced by gD−/−stocks but not by gD−/+stocks. The drug also rescues gD−/−from degradation. (iii) Cells transduced with cation-independent mannose 6-phosphate receptor (CI-MPR) block apoptosis induced by either gD−/−or gD−/+virus. (iv) Expression of sequences antisense to the cloned CI-MPR gene induced apoptosis by themselves. Wild-type virus but not gD−/−or gD−/+stocks of mutant virus blocked apoptosis induced by the expression of CI-MPR antisense sequences. These results exclude the possibility that to block apoptosis, gD must interact with its HveA receptor, a member of the tumor necrosis factor alpha receptor family. Instead, the data suggest that gD blocks the influx of lysosomal enzymes into the endosomal compartment by binding to CI-MPR. This conclusion is consistent with published reports that phosphorylated gD interacts with CI-MPR.