Related Experiment Video
Updated: Jul 16, 2026
Preparation of Cell-lines for Conditional Knockdown of Gene Expression and Measurement of the Knockdown Effects on E4orf4-Induced Cell Death
Published on: October 21, 2012
Apoptosis induction by avian reovirus through p53 and mitochondria-mediated pathway
Julius L C Chulu1, Long H Lee, Ya C Lee
1Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung, Taiwan.
Abstract:
Although induction of apoptosis by avian reovirus has been demonstrated in primary chicken embryonic fibroblast and several cell lines, to date, the potential significance of avian reovirus (ARV)-induced apoptosis and its pathways in cultured cells are still largely unknown. We now provide the first evidence of upregulation of p53 and Bax and specifically for Bax translocation from cytosol to mitochondria following infection with a cytoplasmically replicating RNA virus. Bax translocation to the mitochondria led to the release of mitochondrial proapoptic factors cytochrome c and Smac/DIABLO from mitochondria to the cytosol, but not the release of apoptosis-inducting factor. Activation of caspases-9 and -3 which cleaves the enzyme poly(ADP-ribose) polymerase in ARV-infected BHK-21 cells was also detected. Internucleosomal DNA cleavage was prevented by caspase inhibitors, further demonstrating that ARV-induced apoptosis was executed through caspase-dependent mechanisms. Stable expression of human bcl-2 in BHK-21 cells not only blocked ARV-induced apoptosis and DNA fragmentation but also reduced the level of infectious virus production and its spread in BHK-21 cells infected with ARV at a low multiplicity of infection. All our data suggest that p53 and the mitochondria-mediated pathway played an important regulatory role in ARV-induced apoptosis in BHK-21 cells. To further study the pathogenesis of ARV infection, a dual-labeling assay was used for the simultaneous detection of cells containing viral antigen and apoptotic cells. Dual-labeling assay revealed that the majority of antigen-expressing cells were not apoptotic. Remarkably, some apoptotic but non-antigen-expressing cells were frequently located in the vicinity of antigen-expressing cells. Syncytium formation in ARV-infected BHK-21 cells undergoing apoptosis, was apparent in large syncytia at late infection times, indicating a correlation between virus replication and apoptosis in cultured cells.
Insights
Avian reovirus (ARV) infection triggers apoptosis in cultured cells via the p53 and mitochondria-mediated pathway, involving Bax translocation and caspase activation. Blocking apoptosis with bcl-2 reduced viral spread, highlighting its role in ARV pathogenesis.
Area of Science:
- Virology
- Cell Biology
- Molecular Biology
Background:
- Avian reovirus (ARV) is known to induce apoptosis, but the specific pathways and significance in cultured cells remain largely uncharacterized.
- Understanding ARV-induced apoptosis is crucial for elucidating viral pathogenesis and developing potential therapeutic strategies.
Purpose of the Study:
- To investigate the molecular mechanisms and pathways involved in ARV-induced apoptosis in cultured cells.
- To determine the role of p53, Bax, caspases, and mitochondria in ARV-mediated cell death.
- To assess the impact of blocking apoptosis on viral replication and spread.
Main Methods:
- Analysis of p53 and Bax expression and localization in ARV-infected BHK-21 cells.
- Detection of mitochondrial apoptotic factor release (cytochrome c, Smac/DIABLO).
- Assay of caspase-9, caspase-3 activation, and poly(ADP-ribose) polymerase cleavage.
- Evaluation of internucleosomal DNA cleavage and its inhibition by caspase inhibitors.
- Assessment of ARV-induced apoptosis and viral spread in BHK-21 cells stably expressing human bcl-2.
- Dual-labeling assay to correlate viral antigen expression with apoptosis.
Main Results:
- ARV infection led to upregulation of p53 and Bax, with Bax translocation from cytosol to mitochondria.
- Mitochondrial release of cytochrome c and Smac/DIABLO, followed by caspase-9 and -3 activation, and poly(ADP-ribose) polymerase cleavage.
- ARV-induced apoptosis and DNA fragmentation were caspase-dependent and inhibited by bcl-2 expression.
- Stable bcl-2 expression reduced infectious virus production and spread.
- Dual-labeling revealed that most antigen-expressing cells were not apoptotic, but apoptotic cells were near antigen-positive cells.
- Syncytium formation correlated with apoptosis in late-stage ARV-infected cells.
Conclusions:
- The p53 and mitochondria-mediated pathway plays a significant role in ARV-induced apoptosis in BHK-21 cells.
- ARV-induced apoptosis is executed through caspase-dependent mechanisms.
- Inhibition of apoptosis by bcl-2 not only blocks cell death but also reduces viral replication and spread, suggesting a link between apoptosis and viral pathogenesis.
Related Concept Videos
01:17The Extrinsic Apoptotic Pathway
01:30Apoptosis
01:31The Intrinsic Apoptotic Pathway
01:22Cellular Injury V: Apoptosis and Autophagy
01:30Overview of Cell Death
Cell death was observed in the early 19th century, but there was no experimental evidence to prove it. In 1842, Carl Vogt first discovered cell death in a metamorphic toad; however, it was not termed ‘cell death.’ Scientists discovered different cell death pathways only in the 20th century...
01:18Autophagic Cell Death
Autophagy and Apoptosis
Autophagy can activate apoptosis. In normal conditions, the autophagy activating protein Beclin-1 and pro-apoptotic...