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The apoptosis pathway triggered by the interferon-induced protein kinase PKR requires the third basic domain,
S B Lee1, D Rodríguez, J R Rodríguez
1Centro Nacional de Biotecnología, CSIC Cantoblanco, Madrid, Spain.
Abstract:
The interferon-induced double-stranded RNA-dependent protein kinase (PKR) is a serine/threonine kinase which exerts antiviral and anticellular functions. The antiviral effect of PKR is mediated by the phosphorylation of the alpha subunit of the translational initiation factor elF-2 alpha, while it is not known whether the anticellular effect is due to phosphorylation of elF-2 alpha, l kappa B, or other unknown substrates. We have previously shown that activation of PKR during infection of cells with a vaccinia virus recombinant expressing the wild-type kinase resulted in a complete inhibition of viral and cellular protein synthesis and in the induction of apoptosis. Here, we report that expression of the human proto-oncogene bcl-2 blocks PKR-induced apoptosis but not PKR-induced inhibition of translation. In addition, PKR-induced apoptosis resulted in a cleavage of the death substrate poly(ADP-ribose) polymerase (PARP). Moreover, induction of apoptosis by PKR was not observed with a mutant lacking the third basic region (aa 234-272). Taken together, these results suggest that the third basic region of PKR is required for PKR-induced apoptosis, the process is initiated upstream of bcl-2 and involves activation of a cellular protease, CPP32, or its family members that cleave PARP.
Insights
The double-stranded RNA-dependent protein kinase (PKR) triggers apoptosis, but bcl-2 blocks this cell death. A specific region of PKR is essential for inducing apoptosis, involving protease activation.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- The interferon-induced protein kinase PKR has known antiviral functions via elF-2 alpha phosphorylation.
- PKR's anticellular effects and their substrates (elF-2 alpha, I kappa B, or others) are not fully understood.
- Previous work showed PKR activation inhibits protein synthesis and induces apoptosis.
Purpose of the Study:
- To investigate the role of bcl-2 in PKR-induced apoptosis.
- To identify the molecular mechanisms underlying PKR-induced apoptosis.
- To determine the specific PKR domain required for apoptosis induction.
Main Methods:
- Expression of human proto-oncogene bcl-2 in PKR-activated cells.
- Analysis of protein synthesis inhibition and apoptosis.
- Examination of poly(ADP-ribose) polymerase (PARP) cleavage.
- Use of a PKR mutant lacking the third basic region (aa 234-272).
Main Results:
- Bcl-2 expression blocked PKR-induced apoptosis but not translation inhibition.
- PKR-induced apoptosis involved cleavage of the PARP substrate.
- A PKR mutant lacking the third basic region did not induce apoptosis.
- Apoptosis induction by PKR was upstream of bcl-2 and involved CPP32 or related proteases.
Conclusions:
- The third basic region of PKR is crucial for inducing apoptosis.
- PKR-mediated apoptosis is initiated upstream of bcl-2.
- PKR-induced apoptosis involves the activation of CPP32 family proteases that cleave PARP.