N-terminus cleavage of bcl-2 by a novel cellular non-ICE cysteine proteinase.
The bcl-2 protein plays an essential role in preventing cell death. Its activity is regulated through association with bcl-2 homologous and nonhomologous proteins and also by serine phosphorylation. We now report that bcl-2 can be proteolytically cleaved towards its N-terminus by a cysteine proteinase present in RL-7 lymphoma cell lysates, yielding a major product of apparent MW 20 kDa, different from the products of bcl-2 cleavage by HIV protease. Moreover, bcl-2 proteins mutated for Asp residues at positions 31 and 34 were efficiently cleaved by RL-7 cell lysates, indicating that this proteolytic activity is distinct from the caspase-3 that cleaves bcl-2 at Asp 34. This bcl-2 cleaving activity is inhibited by E-64 and is therefore distinct from the proteinases of the ICE/Ced-3 family (caspases), whereas reciprocally, ICE (caspase-1) is unable to cleave bcl-2. It is optimally active at pH 5, a feature distinguishing it from calpain, another non-ICE cysteine proteinase which has been associated with apoptosis. This novel bcl-2 cleaving protease, although constitutively present in RL-7 cells and resting peripheral blood lymphocytes (PBL) was upregulated following induction of apoptosis in RL-7 cells or mitogen activation in PBL. The N-terminus of bcl-2 which contains the BH4 domain that binds the kinase Raf-1 and the phosphatase calcineurin is essential for anti-apoptotic activity. Its cleavage might provide a novel post-translational mechanism for regulating bcl-2 function and could amplify ongoing programmed cell death.
Yamamoto
A M
AM
INSERM U25, Hôpital Necker, Paris, France.
Eloy
L
L
Bach
J F
JF
Garchon
H J
HJ
D017209:Apoptosis; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D006801:Humans; D016393:Lymphoma, B-Cell / Q000201:enzymology; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism*; D010449:Peptide Mapping; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
D015853:Cysteine Proteinase Inhibitors; D010446:Peptide Fragments; D019253:Proto-Oncogene Proteins c-bcl-2; D003546:Cysteine Endopeptidases