Cleavage and nuclear translocation of the caspase 3 substrate Rho GDP-dissociation inhibitor, D4-GDI, during apoptosis.
While investigating endonucleases potentially involved in apoptosis, an antisera was raised to bovine deoxyribonuclease II, but it recognized a smaller protein of 26 kDa protein in a variety of cell lines. The 26 kDa protein underwent proteolytic cleavage to 22 kDa concomitantly with DNA digestion in cells induced to undergo apoptosis. Sequencing of the 26 kDa protein identified it as the Rho GDP-dissociation inhibitor D4-GDI. Zinc, okadaic acid, calyculin A, cantharidin, and the caspase inhibitor z-VAD-fmk, all prevented the cleavage of D4-GDI, DNA digestion, and apoptosis. The 26 kDa protein resided in the cytoplasm of undamaged cells, whereas following cleavage, the 22 kDa form translocated to the nucleus. Human D4-GDI, and D4-GDI mutated at the caspase 1 or caspase 3 sites, were expressed in Chinese hamster ovary cells which show no detectable endogenous D4-GDI. Mutation at the caspase 3 site prevented D4-GDI cleavage but did not inhibit apoptosis induced by staurosporine. The cleavage of D4-GDI could lead to activation of Jun N-terminal kinase which has been implicated as an upstream regulator of apoptosis in some systems. However, the results show that the cleavage of D4-GDI and translocation to the nucleus do not impact on the demise of the cell.
Krieser
R J
RJ
Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, New Hampshire 03755, USA.
Eastman
A
A
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001692:Biological Transport; D016466:CHO Cells; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D002417:Cattle; D002467:Cell Nucleus; D006224:Cricetinae; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020730:Guanine Nucleotide Dissociation Inhibitors; D006153:Guanosine Diphosphate / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D014162:Transfection; D062949:rho Guanine Nucleotide Dissociation Inhibitor beta; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; D020743:rhoB GTP-Binding Protein
C569658:ARHGDIB protein, human; D020730:Guanine Nucleotide Dissociation Inhibitors; D008565:Membrane Proteins; D062949:rho Guanine Nucleotide Dissociation Inhibitor beta; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; D006153:Guanosine Diphosphate; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases; D019204:GTP-Binding Proteins; D020743:rhoB GTP-Binding Protein