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Signal-induced site specific phosphorylation targets Bcl2 to the proteasome pathway
1Rammelkamp Center for Education and Research, Department of Pharmacology, Ireland Cancer Center, MetroHealth/Case Western Reserve University, 2500 MetroHealth Drive, Cleveland, OH 44109-1998, USA.
Abstract:
The oncogene derived protein Bcl2 and its family members such as Bcl-xL or Mcl-1 can confer negative control in the pathway of cellular suicide machinery. The reversible phosphorylation of the components in the apoptotic-signaling pathway is likely to be an important regulatory mechanism to control the fate of a cell. Previous reports by others and us demonstrate that phosphorylation of anti-apoptotic proteins such as Bcl2, Bcl-xL or Mcl-1 can regulate their function depending on the apoptotic trigger or cell type. Also, evidence is now accumulating that the ubiquitin proteasome pathway can play an important role in apoptosis. In order to understand whether any cross-talk exists between proteasome and Bcl2 phosphorylation pathways, studies were undertaken employing cell permeable proteasome inhibitors. When proteasomes were inactivated, enhanced accumulation of slower mobility forms of Bcl2 was clearly evident. Due to substitution of the major phosphorylation sites Ser 70, 87 to Ala, no such effect was observed. It is known that in contrary to phospho Bcl2, native Bcl2 (non-phosphoform) is unable to associate with cis-trans peptidyl prolyl isomerase Pin1-a key factor to regulate the fate of phosphoforms of Bcl2 and apoptosis. Thus the enhanced resistance to cell death exhibited by phosphorylation defective mutant Bcl2 might be attributed to its inability to associate with Pin1.
Insights
The proteasome pathway influences Bcl2 phosphorylation, affecting cell death. Blocking proteasomes increases phosphorylated Bcl2, impacting apoptosis regulation and Pin1 interaction.
Area of Science:
- Cellular Biology
- Molecular Biology
- Biochemistry
Background:
- The Bcl2 protein family regulates apoptosis (programmed cell death).
- Phosphorylation and the ubiquitin proteasome system are key regulators of apoptosis.
- Potential cross-talk between Bcl2 phosphorylation and proteasome activity is not fully understood.
Purpose of the Study:
- To investigate the interplay between the proteasome pathway and Bcl2 phosphorylation.
- To determine if proteasome inhibition affects Bcl2 phosphorylation status.
- To explore the role of phosphorylation sites and Pin1 interaction in this process.
Main Methods:
- Utilized cell-permeable proteasome inhibitors to block proteasome activity.
- Analyzed Bcl2 phosphorylation by examining mobility shifts in protein gels.
- Employed phosphorylation-defective Bcl2 mutants (Ser70, 87 to Ala) for functional studies.
- Assessed the interaction of Bcl2 forms with Pin1 (a peptidyl prolyl isomerase).
Main Results:
- Proteasome inhibition led to increased accumulation of slower-mobility (phosphorylated) forms of Bcl2.
- This effect was abolished in Bcl2 mutants lacking major phosphorylation sites.
- Phosphorylation-defective Bcl2 mutants showed reduced association with Pin1.
- This suggests a mechanism linking proteasome activity, Bcl2 phosphorylation, and Pin1 interaction.
Conclusions:
- The ubiquitin proteasome pathway modulates Bcl2 phosphorylation.
- Phosphorylation of Bcl2 is crucial for its interaction with Pin1, influencing cell fate.
- Dysregulation of this cross-talk may contribute to altered apoptosis resistance.