Caspase-7 gene disruption reveals an involvement of the enzyme during the early stages of apoptosis.
Caspases play a key role during apoptotic execution. In an attempt to elucidate the specific role of caspase-7 we generated a chicken DT40 cell line in which both alleles of the gene were disrupted. Viability assays showed that caspase-7-/- clones are more resistant to the common apoptosis-inducing drugs etoposide and staurosporine. Caspase-7-/- cells show a delay in phosphatidylserine externalization and DNA fragmentation as well as cleavage of the caspase substrates poly(ADP-ribose) polymerase 1 and lamins B1 and B2. Caspase affinity labeling and activity assays indicated that deficient cells exhibit a delay in caspase activation compared with wild type DT40 cells, providing an explanation for the differences in apoptotic execution between caspase-7 null and wild type DT40 cells. These results strongly suggest that caspase-7 is involved earlier than other effector caspases in the apoptotic execution process in DT40 B lymphocytes.
Wellcome Trust Centre for Cell Biology, University of Edinburgh, Mayfield Road, Edinburgh EH9 3JR, Scotland, United Kingdom.
Nadia
N
Korfali
Sandrine
S
Ruchaud
David
D
Loegering
Delphine
D
Bernard
Colin
C
Dingwall
Scott H
SH
Kaufmann
William C
WC
Earnshaw
D000818:Animals; D030801:Animals, Genetically Modified; D017304:Annexin A5 / Q000494:pharmacology; D017209:Apoptosis; D015139:Blotting, Southern; D053179:Caspase 7; D020169:Caspases / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002470:Cell Survival; D002645:Chickens; D004396:Coloring Agents / Q000494:pharmacology; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D005047:Etoposide / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D020287:In Situ Nick-End Labeling; D007700:Kinetics; D034921:Lamin Type B / Q000737:chemistry; D008957:Models, Genetic; D010718:Phosphatidylserines / Q000737:chemistry; D011065:Poly(ADP-ribose) Polymerases / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019311:Staurosporine / Q000494:pharmacology; D013997:Time Factors
D017304:Annexin A5; D004396:Coloring Agents; D034921:Lamin Type B; D010718:Phosphatidylserines; D012333:RNA, Messenger; D005047:Etoposide; D011065:Poly(ADP-ribose) Polymerases; C505520:CASP7 protein, human; D053179:Caspase 7; D020169:Caspases; D019311:Staurosporine