Apoptosis is associated with cleavage of a 5 kDa fragment from RB which mimics dephosphorylation and modulates E2F binding.
Dephosphorylation of the RB protein has been reported to be associated with apoptosis. In contrast, we show that treatment of HL60 cells with etoposide or cytosine arabinoside or treatment of breast epithelial cells with alpha-FAS is associated with the cleavage of a 5 kDa fragment from the C-terminus of RB, resulting in a truncated product that we have designated as p100cl. This cleavage event coincides with the activation of cysteine proteases at the onset of apoptosis, is blocked by the addition of iodoacetamide to cells prior to the onset of apoptosis, and results in the expression of faster migrating protein species which can mimic dephosphorylated RB. The free 5 kDa fragment is detected only during apoptosis, predicts a cleavage site that we have mapped to a unique CPP32-like recognition sequence which is present at the C-terminus of all reported RB homologues, and results in a truncated RB protein with enhanced E2F binding affinity. While the causality for this cleavage event in the apoptotic process is still under investigation, our findings suggest distinct post-translational pathways for the RB product between cells examined during growth arrest (p105 hypophosphorylated RB) or apoptosis (p100cl).
Chen
W D
WD
NCI-Navy Medical Oncology Branch, Division of Clinical Sciences, National Cancer Institute, Bethesda, Maryland 20889, USA.
Otterson
G A
GA
Lipkowitz
S
S
Khleif
S N
SN
Coxon
A B
AB
Kaye
F J
FJ
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D001943:Breast Neoplasms / Q000188:drug therapy / Q000378:metabolism; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D003546:Cysteine Endopeptidases / Q000378:metabolism; D003561:Cytarabine / Q000494:pharmacology; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D005047:Etoposide / Q000494:pharmacology; D005260:Female; D018922:HL-60 Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D010446:Peptide Fragments / Q000378:metabolism*; D010766:Phosphorylation; D016160:Retinoblastoma Protein / Q000378:metabolism*; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000378:metabolism*; D019014:fas Receptor / Q000502:physiology
D000970:Antineoplastic Agents; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D010446:Peptide Fragments; D016160:Retinoblastoma Protein; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors; D019014:fas Receptor; D003561:Cytarabine; D005047:Etoposide; D003546:Cysteine Endopeptidases