Type I cAMP-dependent protein kinase delays apoptosis in human neutrophils at a site upstream of caspase-3.
Current data suggest that apoptosis controls neutrophil numbers in tissues. We analyzed roles for and the sites of action for the cAMP-dependent protein kinases (cAPKs) in apoptosis induced in human neutrophils by in vitro storage, cycloheximide (CHX) exposure, and anti-Fas exposure. Treatment with 8-chlorophenylthio-cAMP (8-CPT-cAMP) prolonged the time required for 50% of the cells to exhibit apoptotic morphology (t50) from 16.3 to 41.8 h (in vitro culture), from 2.4 to 7.8 h (CHX), and from 4.8 to 6.5 h (anti-Fas). CHX +/- 8-CPT-cAMP did not significantly alter resting intracellular calcium levels and H-89, a selective inhibitor of cAPK, had no effect on apoptosis in the absence of the analogue. In contrast, site-selective cAMP analogues that specifically activated the type I cAPK, but not type II cAPK, synergistically attenuated apoptosis. Exposure to 8-CPT-cAMP delayed, in parallel, the activity of caspase-3 (CPP-32beta), whereas mitogen-activated protein kinase kinase (MAPKK) inhibitor, PD98059, had no effect on CHX-induced apoptosis +/- 8-CPT-cAMP. Together these results indicate that type I cAPK activation is necessary and sufficient to mediate cAMP-induced delay in human neutrophil apoptosis induced by several mechanisms and suggest that one of the major sites of cAPK action is upstream of caspase-3 (CPP-32beta) activation.
Parvathenani
L K
LK
Center for Pediatric Research, Children's Hospital of The King's Daughters, Eastern Virginia Medical School, Norfolk, Virginia 23510, USA.
Buescher
E S
ES
Chacon-Cruz
E
E
Beebe
S J
SJ
D000328:Adult; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003513:Cycloheximide / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004789:Enzyme Activation; D005260:Female; D006801:Humans; D008297:Male; D009504:Neutrophils / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D013873:Thionucleotides / Q000494:pharmacology
D013873:Thionucleotides; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D003513:Cycloheximide; D000242:Cyclic AMP; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; D002118:Calcium