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Double-stranded RNA is a trigger for apoptosis in vaccinia virus-infected cells
K V Kibler1, T Shors, K B Perkins
1Department of Microbiology, Arizona State University, Tempe 85287-2701, USA.
Abstract:
The vaccinia virus E3L gene codes for double-stranded RNA (dsRNA) binding proteins which can prevent activation of the dsRNA-dependent, interferon-induced protein kinase PKR. Activated PKR has been shown to induce apoptosis in HeLa cells. HeLa cells infected with vaccinia virus with the E3L gene deleted have also been shown to undergo apoptosis, whereas HeLa cells infected with wild-type vaccinia virus do not. In this report, using virus recombinants expressing mutant E3L products or alternative dsRNA binding proteins, we show that suppression of induction of apoptosis correlates with functional binding of proteins to dsRNA. Infection of HeLa cells with ts23, which leads to synthesis of increased dsRNA at restrictive temperature, induced apoptosis at restrictive but not permissive temperatures. Treatment of cells with cytosine arabinoside, which blocks the late buildup of dsRNA in vaccinia virus-infected cells, prevented induction of apoptosis by vaccinia virus with E3L deleted. Cells transfected with dsRNA in the absence of virus infection also underwent apoptosis. These results suggest that dsRNA is a trigger that can initiate a suicide response in virus-infected and perhaps uninfected cells.
Insights
Double-stranded RNA (dsRNA) triggers apoptosis, a cellular suicide response. Vaccinia virus E3L protein binding to dsRNA prevents this, protecting cells from viral infection-induced cell death.
Area of Science:
- Virology
- Molecular Biology
- Immunology
Background:
- The vaccinia virus E3L gene encodes proteins that bind double-stranded RNA (dsRNA).
- This binding prevents activation of the dsRNA-dependent protein kinase PKR.
- Activated PKR can induce apoptosis (programmed cell death) in host cells.
Purpose of the Study:
- To investigate the role of dsRNA binding proteins in preventing apoptosis during vaccinia virus infection.
- To determine if dsRNA itself acts as a trigger for apoptosis.
Main Methods:
- Utilized vaccinia virus recombinants expressing mutant E3L proteins or alternative dsRNA binding proteins.
- Infected HeLa cells with modified vaccinia viruses and observed apoptosis induction.
- Used temperature-sensitive mutants (ts23) and cytosine arabinoside treatment to manipulate dsRNA levels.
- Transfected cells with dsRNA in the absence of viral infection.
Main Results:
- Suppression of apoptosis correlated with the functional binding of proteins to dsRNA.
- Increased dsRNA synthesis at restrictive temperatures induced apoptosis in ts23-infected cells.
- Blocking dsRNA buildup with cytosine arabinoside prevented apoptosis in E3L-deleted virus-infected cells.
- Transfection with dsRNA alone induced apoptosis in uninfected cells.
Conclusions:
- Functional binding of dsRNA by viral proteins is crucial for preventing apoptosis.
- Double-stranded RNA serves as a trigger for initiating apoptosis.
- This apoptotic response can occur in both virus-infected and uninfected cells.