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Induction of apoptosis by Sindbis virus occurs at cell entry and does not require virus replication
1Department of Molecular Microbiology, Johns Hopkins University School of Hygiene and Public Health, Baltimore, Maryland 21205, USA.
Abstract:
Sindbis virus (SV) is an alphavirus that causes encephalitis in mice and can lead to the apoptotic death of infected cells. To determine the step in virus replication during which apoptosis is triggered, we used UV-inactivated SV, chemicals that block virus fusion or protein synthesis, and cells that do and do not express heparan sulfate, the initial binding molecule for SV infection of many cells. In initial experiments, UV-inactivated neuroadapted SV (NSV) induced apoptosis in Chinese hamster ovary (CHO) cells lacking heparan sulfate in the presence of cycloheximide. When fusion of prebound UV-inactivated NSV was rapidly induced at the plasma membrane by exposure to acidic pH, apoptosis was induced in CHO cells with or without heparan sulfate in the presence or absence of cycloheximide in a virus dose-dependent manner. In N18 neuroblastoma cells, the relative virulence of the virus strain was an important determinant of apoptosis induced by UV-inactivated SV. Treatment of N18 cells with monensin to prevent endosomal acidification an hour before, but not 2 h after, exposure to live NSV blocked the induction of cell death, as did treatment with NH(4)Cl or bafilomycin A1. These studies indicate that SV can induce apoptosis at the time of fusion with the cell membrane and that virus replication is not required.
Insights
Sindbis virus (SV) triggers cell death (apoptosis) during membrane fusion, not requiring viral replication. This alphavirus-induced apoptosis occurs early in infection, even with inactivated virus.
Area of Science:
- Virology
- Cell Biology
- Immunology
Background:
- Sindbis virus (SV), an alphavirus, is known to cause encephalitis and induce apoptotic cell death.
- Understanding the precise stage of viral replication that triggers apoptosis is crucial for developing targeted antiviral strategies.
Purpose of the Study:
- To pinpoint the specific step in Sindbis virus replication that initiates apoptosis in host cells.
- To investigate the role of viral fusion and replication in triggering SV-induced apoptosis.
Main Methods:
- Utilized UV-inactivated SV, fusion inhibitors, and protein synthesis blockers.
- Employed Chinese hamster ovary (CHO) and N18 neuroblastoma cells, with and without heparan sulfate expression.
- Manipulated endosomal acidification using monensin, NH(4)Cl, and bafilomycin A1.
Main Results:
- UV-inactivated SV induced apoptosis upon fusion with the cell membrane, independent of heparan sulfate presence.
- Apoptosis induction was virus dose-dependent and occurred even without protein synthesis.
- Blocking endosomal acidification prevented apoptosis in live SV-infected cells, indicating fusion is a key trigger.
Conclusions:
- Sindbis virus can induce apoptosis during the membrane fusion process.
- Viral replication is not a prerequisite for triggering SV-induced apoptosis.
- The timing of fusion and endosomal acidification are critical factors in SV-mediated cell death.