Related Experiment Videos
Inactivation of the inhibitory kappaB protein kinase/nuclear factor kappaB pathway by Par-4 expression potentiates
M T Diaz-Meco1, M J Lallena, A Monjas
1Laboratorio Glaxo Wellcome-CSIC de Biología Molecular y Celular, Centro de Biología Molecular "Severo Ochoa" (Consejo Superior de Investigaciones Científicas-Universidad Autónoma de Madrid), Spain.
Abstract:
Par-4 is a novel protein identified in cells undergoing apoptosis. The ability of Par-4 to promote apoptotic cell death is dependent on the binding and inactivation of the atypical protein kinases C (PKCs). This subfamily of kinases has been reported to control nuclear factor kappaB (NF-kappaB) through the regulation of the IkappaB kinase activity. NF-kappaB activation by tumor necrosis factor alpha (TNFalpha) provides a survival signal that impairs the TNFalpha-induced apoptotic response. We show here that expression of Par-4 inhibits the TNFalpha-induced nuclear translocation of p65 as well as the kappaB-dependent promoter activity. Interestingly, Par-4 expression blocks inhibitory kappaB protein (IkappaB) kinase activity, which leads to the inhibition of IkappaB phosphorylation and degradation, in a manner that is dependent on its ability to inhibit lambda/iotaPKC. Of potential functional relevance, the expression of Par-4 allows TNFalpha to induce apoptosis in NIH-3T3 cells. In addition, the down-regulation of Par-4 levels by oncogenic Ras sensitizes cells to TNFalpha-induced NF-kappaB activation.
Insights
Par-4 protein promotes apoptosis by inhibiting atypical protein kinases C (PKCs). This blocks NF-kappaB activation, enhancing TNFalpha-induced cell death and overcoming survival signals.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Apoptosis is a critical cellular process.
- Atypical protein kinases C (PKCs) and nuclear factor kappaB (NF-kappaB) signaling pathways regulate cell survival and death.
- Tumor necrosis factor alpha (TNFalpha) signaling can induce apoptosis but is often counteracted by survival pathways.
Purpose of the Study:
- To investigate the role of Par-4 in apoptosis.
- To determine the mechanism by which Par-4 influences NF-kappaB signaling.
- To assess the functional consequences of Par-4 expression on TNFalpha-induced apoptosis.
Main Methods:
- Expression of Par-4 in NIH-3T3 cells.
- Analysis of p65 nuclear translocation.
- Measurement of kappaB-dependent promoter activity.
- Assay of inhibitory kappaB protein (IkappaB) kinase activity.
- Assessment of IkappaB phosphorylation and degradation.
- Evaluation of TNFalpha-induced apoptosis.
Main Results:
- Par-4 expression inhibited TNFalpha-induced nuclear translocation of p65 and kappaB-dependent promoter activity.
- Par-4 blocked IkappaB kinase activity, preventing IkappaB phosphorylation and degradation.
- This inhibition was dependent on Par-4's ability to inhibit lambda/iotaPKC.
- Par-4 expression sensitized NIH-3T3 cells to TNFalpha-induced apoptosis.
- Oncogenic Ras-mediated down-regulation of Par-4 sensitized cells to TNFalpha-induced NF-kappaB activation.
Conclusions:
- Par-4 promotes apoptosis by inhibiting atypical PKCs and consequently blocking NF-kappaB activation.
- Par-4 sensitizes cells to TNFalpha-induced apoptosis by disrupting survival signaling.
- These findings elucidate a novel mechanism linking Par-4, PKCs, and NF-kappaB in the regulation of cell death.