Dual effects of staurosporine on A431 and NRK cells: microfilament disassembly and uncoordinated lamellipodial activity followed by cell death.
The general protein kinase inhibitor staurosporine (STS) has dual effects on human epidermoid cancer cells (A431) and normal rat kidney fibroblasts (NRK). It almost immediately stimulated increased lamellipodial activity of both cell lines and after 2 h induced typical signs of apoptosis, including cytoplasmic condensation, nuclear fragmentation, caspase-3 activation and DNA degradation. In the early phase we observed disruption of actin-containing stress fibres and accumulation of monomeric actin in the perinuclear region and cell nucleus. Increased lamellipodial-like extensions were observed particularly in A431 cells as demonstrated by co-localisation of actin and Arp2/3 complex, whereas NRK cells shrunk and exhibited numerous thin long extensions. These extensions exhibited uncoordinated centrifugal motile activity that appeared to tear the cells apart. Both cofilin and ADF were translocated from perinuclear regions to the cell cortex and, as expected in the presence of a kinase inhibitor, all the cofilin was dephosphorylated. Myosin II was absent from the extensions, and a reduction of phosphorylated myosin light chains was observed within the cytoplasm indicating myosin inactivation. Microtubules and intermediate filaments retained their characteristic filamentous organisation after STS exposure even when the cells became rounded and disorganised. Simultaneous treatment of NRK cells with STS and the caspase inhibitor zVAD did not inhibit the morphological and cytoskeletal changes. However, the cells underwent cell death as verified by positive annexin-V-staining. Thus it seems likely that cell death induced by STS may not only be a consequence of the activation of caspase, instead the disruption of the many motile processes involving the actin cytoskeleton may by itself suffice to induce caspase-independent cell death.
Mannherz
Hans G
HG
Department of Anatomy and Embryology, Ruhr-University, Bochum, Germany. hans.mannherz@ruhr-uni-bochum.de
Gonsior
Sabine M
SM
Wu
Xueqing
X
Polzar
Bernhard
B
Pope
Brian J
BJ
Wartosch
Lena
L
Weeds
Alan G
AG
D008841:Actin Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D051339:Actin Depolymerizing Factors / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D046529:Microscopy, Electron, Transmission; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D024462:Myosin Type II / Q000378:metabolism / Q000502:physiology; D011554:Pseudopodia / Q000187:drug effects* / Q000502:physiology; D019311:Staurosporine / Q000494:pharmacology*
D051339:Actin Depolymerizing Factors; D000199:Actins; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D020169:Caspases; D024462:Myosin Type II; D019311:Staurosporine