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CREB activation induced by mitochondrial dysfunction is a new signaling pathway that impairs cell proliferation
T Arnould1, S Vankoningsloo, P Renard
1Laboratoire de Biochimie et Biologie Cellulaire, University of Namur (FUNDP), 61 rue de Bruxelles, B-5000 Namur, Belgium. thierry.arnould@fundp.ac.be
Abstract:
We characterized a new signaling pathway leading to the activation of cAMP-responsive element-binding protein (CREB) in several cell lines affected by mitochondrial dysfunction. In vitro kinase assays, inhibitors of several kinase pathways and overexpression of a dominant-negative mutant for calcium/calmodulin kinase IV (CaMKIV), which blocks the activation of CREB, showed that CaMKIV is activated by a mitochondrial activity impairment. A high calcium concentration leading to the disruption of the protein interaction with protein phosphatase 2A explains CaMKIV activation in these conditions. Transcrip tionally active phosphorylated CREB was also found in a rho0 143B human osteosarcoma cell line and in a MERRF cybrid cell line mutated for tRNA(Lys) (A8344G). We also showed that phosphorylated CREB is involved in the proliferation defect induced by a mitochondrial dysfunction. Indeed, cell proliferation inhibition can be prevented by CaMKIV inhibition and CREB dominant-negative mutants. Finally, our data suggest that phosphorylated CREB recruits p53 tumor suppressor protein, modifies its transcriptional activity and increases the expression of p21(Waf1/Cip1), a p53-regulated cyclin-dependent kinase inhibitor.
Insights
Mitochondrial dysfunction activates calcium/calmodulin kinase IV (CaMKIV), leading to cAMP-responsive element-binding protein (CREB) activation. This phosphorylated CREB impairs cell proliferation and interacts with p53, impacting cell cycle regulation.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Mitochondrial dysfunction is implicated in various diseases.
- The signaling pathways linking mitochondrial dysfunction to cellular responses are not fully understood.
- cAMP-responsive element-binding protein (CREB) is a transcription factor involved in cellular processes.
Purpose of the Study:
- To elucidate the signaling pathway activated by mitochondrial dysfunction leading to CREB activation.
- To investigate the role of CREB in the cellular defects associated with mitochondrial dysfunction.
- To explore the interaction between CREB and other cellular regulators like p53.
Main Methods:
- In vitro kinase assays and pathway inhibitors were used to identify key kinases.
- Dominant-negative mutants were employed to block specific signaling pathways.
- Cell lines with induced mitochondrial dysfunction (rho0 and MERRF cybrid) were utilized.
- Western blotting and co-immunoprecipitation were used to assess protein interactions and modifications.
Main Results:
- Mitochondrial activity impairment activates calcium/calmodulin kinase IV (CaMKIV).
- CaMKIV activation is mediated by high intracellular calcium disrupting protein phosphatase 2A interaction.
- Phosphorylated CREB is present in cells with mitochondrial defects and contributes to proliferation inhibition.
- Inhibition of CaMKIV or CREB rescues proliferation defects.
- Phosphorylated CREB interacts with p53, increasing p21(Waf1/Cip1) expression.
Conclusions:
- A novel signaling cascade links mitochondrial dysfunction to CREB activation via CaMKIV.
- Activated CREB plays a critical role in mediating the proliferation defects caused by mitochondrial dysfunction.
- The CREB-p53 interaction represents a new mechanism by which mitochondrial dysfunction impacts cell cycle control through p21(Waf1/Cip1) upregulation.