The catalytic domain of xPAK1 is sufficient to induce myosin II dependent in vivo cell fragmentation independently of other apoptotic events.
During apoptosis, cells are fragmented into sealed packages for safe disposal by phagocytosis, a process requiring major reorganisation of the cytoskeleton. The small p21 GTPase-activated kinases (PAKs) have been implicated in regulating cytoskeletal dynamics and a subset are activated by caspase 3/7 cleavage. However, the functional importance of this activation in apoptosis remains unknown. Using early Xenopus embryos, we have dissected xPAK1 activation from other causative events in apoptosis. An apoptotic-like cell fragmentation was observed 30 min after expression of the xPAK1 catalytic domain and occurred in the absence of other markers of apoptosis. In vitro, activated xPAK1 phosphorylated the regulatory light chain (xMLC) of myosin II at threonine 18 and serine 19, events known to activate the actin-dependent ATPase of cytoskeletal myosin. In vivo, activated xPAK1 induced hyperphosphorylation of xMLC. BDM, a myosin inhibitor, and ML-7, a MLCK inhibitor, both abrogated cell fragmentation induced by activated xPAK1, and ML-7 also inhibited xPAK1 activity. Endogenous xPAK1 was cleaved during normal apoptosis and this was associated with xPAK1 activation and increased serine 19 phosphorylation of xMLC. The data show that PAK activation is sufficient for apoptotic body formation in vivo and strongly suggest that activation of myosin II is essential for this process.
Bisson
Nicolas
N
Cancer Research Centre and Department of Medical Biology, Faculty of Medicine, Laval University, Hôtel-Dieu de Québec, 9 rue McMahon, Québec, QC, G1R 2J6, Canada.
Islam
Nazrul
N
Poitras
Luc
L
Jean
Steve
S
Bresnick
Anne
A
Moss
Tom
T
D000199:Actins / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D001381:Azepines / Q000494:pharmacology; D001757:Blastomeres / Q000166:cytology; D020134:Catalytic Domain; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology; D024462:Myosin Type II / Q000502:physiology*; D009219:Myosin-Light-Chain Kinase / Q000502:physiology; D009281:Naphthalenes / Q000494:pharmacology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000502:physiology*; D014981:Xenopus / Q000196:embryology*; D054462:p21-Activated Kinases
D000199:Actins; D001381:Azepines; D009281:Naphthalenes; C070571:ML 7; D017346:Protein Serine-Threonine Kinases; D054462:p21-Activated Kinases; D009219:Myosin-Light-Chain Kinase; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D024462:Myosin Type II