APOPTOSIS IS ASSOCIATED WITH CLEAVAGE OF A 5 KDA FRAGMENT FROM RB WHICH MIMICS DEPHOSPHORYLATION AND MODULATES E2F BINDING

Dephosphorylation of the RE protein has been reported to be associated with apoptosis, In contrast, we show that treatment of HL60 cells wit h etoposide or cytosine arabinoside or treatment of breast epithelial cells with alpha-FAS is associated with the cleavage of a 5 kDa fragme nt from the C-terminus of RE, resulting in a truncated product that we have designated as p100cl. This cleavage event coincides with the act ivation of cysteine proteases at the onset of apoptosis, is blocked by the addition of iodoacetamide to cells prior to the onset of apoptosi s, and results in the expression of faster migrating protein species w hich can mimic dephosphorylated RE. The free 5 kDa fragment is detecte d 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 RE homologues, and results in a truncated RE protein with enhanced E2F binding affinity, While the causality for th is cleavage event in the apoptotic process is still under investigatio n, our findings suggest distinct post-translational pathways for the R E product between cells examined during growth arrest (p105 hypophosph orylated RE) or apoptosis (p100cl).