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p28 Bap31, a Bcl-2/Bcl-XL- and procaspase-8-associated protein in the endoplasmic reticulum
1Department of Biochemistry, McIntyre Medical Sciences Building, McGill University, Montreal, Quebec, Canada H3G 1Y6.
Abstract:
We have identified a human Bcl-2-interacting protein, p28 Bap31. It is a 28-kD (p28) polytopic integral protein of the endoplasmic reticulum whose COOH-terminal cytosolic region contains overlapping predicted leucine zipper and weak death effector homology domains, flanked on either side by identical caspase recognition sites. In cotransfected 293T cells, p28 is part of a complex that includes Bcl-2/Bcl-XL and procaspase-8 (pro-FLICE). Bax, a pro-apoptotic member of the Bcl-2 family, does not associate with the complex; however, it prevents Bcl-2 from doing so. In the absence (but not presence) of elevated Bcl-2 levels, apoptotic signaling by adenovirus E1A oncoproteins promote cleavage of p28 at the two caspase recognition sites. Purified caspase-8 (FLICE/MACH/Mch5) and caspase-1(ICE), but not caspase-3 (CPP32/apopain/ Yama), efficiently catalyze this reaction in vitro. The resulting NH2-terminal p20 fragment induces apoptosis when expressed ectopically in otherwise normal cells. Taken together, the results suggest that p28 Bap31 is part of a complex in the endoplasmic reticulum that mechanically bridges an apoptosis-initiating caspase, like procaspase-8, with the anti-apoptotic regulator Bcl-2 or Bcl-XL. This raises the possibility that the p28 complex contributes to the regulation of procaspase-8 or a related caspase in response to E1A, dependent on the status of the Bcl-2 setpoint within the complex.
Insights
Researchers identified p28 Bap31, an endoplasmic reticulum protein that bridges apoptosis regulators like Bcl-2 and procaspase-8. Cleavage of p28 Bap31 by caspases can trigger apoptosis, revealing a new regulatory mechanism.
Area of Science:
- Cellular biology
- Molecular biology
- Apoptosis research
Background:
- Bcl-2 family proteins regulate apoptosis.
- Caspases are key executioners of apoptosis.
- Endoplasmic reticulum proteins play roles in cell death pathways.
Purpose of the Study:
- To identify and characterize novel human proteins interacting with Bcl-2.
- To elucidate the role of p28 Bap31 in apoptosis regulation.
- To investigate the interplay between p28 Bap31, Bcl-2, and caspases.
Main Methods:
- Co-transfection of 293T cells.
- Protein complex analysis.
- In vitro caspase cleavage assays.
- Ectopic expression of protein fragments.
Main Results:
- Identified p28 Bap31, an endoplasmic reticulum protein with caspase recognition sites.
- p28 Bap31 forms a complex with Bcl-2/Bcl-XL and procaspase-8.
- Adenovirus E1A signaling induces p28 Bap31 cleavage by caspase-8 or caspase-1.
- The N-terminal p20 fragment of cleaved p28 Bap31 induces apoptosis.
Conclusions:
- p28 Bap31 acts as a scaffold, linking procaspase-8 and Bcl-2/Bcl-XL at the endoplasmic reticulum.
- Cleavage of p28 Bap31 by caspases is a critical step in E1A-induced apoptosis.
- The p28 Bap31 complex offers a novel regulatory point for apoptosis control.