Tissue transglutaminase-dependent posttranslational modification of the retinoblastoma gene product in promonocytic cells undergoing apoptosis.
The retinoblastoma gene product (pRB) plays an important role in controlling both cell release from the G1 phase and apoptosis. We show here that in the early phases of apoptosis, pRB is posttranslationally modified by a tissue transglutaminase (tTG)-catalyzed reaction. In fact, by employing a novel haptenized lysis synthetic substrate which allows the isolation of glutaminyl-tTG substrates in vivo, we identified pRB as a potential tTG substrate in U937 cells undergoing apoptosis. In keeping with this finding, we showed that apoptosis of U937 cells is characterized by the rapid disappearance of the 105,000- to 110,000-molecular-weight pRB forms concomitantly with the appearance of a smear of immunoreactive products with a molecular weight of greater than 250,000. The shift in pRB molecular weight was reproduced by adding exogenous purified tTG to extracts obtained from viable U937 cells and was prevented by dansylcadaverine, a potent enzyme inhibitor. The effect of the pRB posttranslational modification during apoptosis was investigated by determining the E2F-1 levels and by isolating and characterizing pRB-null clones from U937 cells. Notably, the lack of pRB in these U937-derived clones renders these p53-null cells highly resistant to apoptosis induced by serum withdrawal, calphostin C, and ceramide. Taken together, these data suggest that tTG, acting on the pRB protein, might play an important role in the cell progression through the death program.
Oliverio
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Department of Biology, University of Rome Tor Vergata, Italy.
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Di Sano
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Farrace
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Fesus
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Nemes
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Piredda
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Spinedi
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Piacentini
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D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001769:Blood; D002103:Cadaverine / Q000031:analogs & derivatives / Q000494:pharmacology; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002460:Cell Line; D002518:Ceramides / Q000494:pharmacology; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019204:GTP-Binding Proteins; D006801:Humans; D009000:Monocytes / Q000166:cytology / Q000378:metabolism*; D009154:Mutation; D009281:Naphthalenes / Q000494:pharmacology; D010766:Phosphorylation; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000032:analysis; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000032:analysis; D011503:Transglutaminases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014212:Tretinoin / Q000494:pharmacology
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002518:Ceramides; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D004791:Enzyme Inhibitors; D009281:Naphthalenes; D019253:Proto-Oncogene Proteins c-bcl-2; D016160:Retinoblastoma Protein; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors; C101954:N-caproylsphingosine; D014212:Tretinoin; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D011493:Protein Kinase C; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C058819:calphostin C; C008542:monodansylcadaverine; D002103:Cadaverine