Marginal blebbing during the early stages of TNF-induced apoptosis indicates alteration in actomyosin contractility.
Dynamics of alterations of cell surface topography during TNF-induced apoptosis of HeLa cells was examined by phase-contrast videomicroscopy and immunomorphological analysis. The final stage of apoptosis accompanied by cell rounding and general blebbing of the cell surface became after 4-6 h of incubation but much earlier, after 1.5-3 h, essentially flattened lamellipodia at the active edges transformed into the small blebs that were continuously extended and retracted during the next 1-2 h. This phenomenon was called "marginal blebbing". It took place before the cytochrome c release from mitochondria to cytosol. Marginal blebbing was inhibited by drugs that depolymerized actin microfilaments (cytochalasin, latrunculin) or decreased Rho-kinase-dependent contractility of actin-myosin cortex (H7, HA-1077, Y27632). A pancaspase inhibitor, zVAD-fmk, completely prevented marginal and general blebbing, and TNF-induced apoptosis. DEVD-fmk, a specific inhibitor of caspase-3, inhibited both marginal and general blebbing but not cell rounding and death. Thus, marginal blebbing is an early microfilament-dependent apoptotic event. It is suggested that it is initiated by minimal activation of caspase-3 and the following local Rho-kinase-dependent stimulation of actin-myosin cortex contractility. Localization of marginal blebs at the active edge may be associated with special organization of cortex in that zone.
Domnina
L V
LV
Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119992, Russia. domina@mbio.genebee.msu.su
Ivanova
O Y
OY
Pletjushkina
O Y
OY
Fetisova
E K
EK
Chernyak
B V
BV
Skulachev
V P
VP
Vasiliev
J M
JM
D008841:Actin Cytoskeleton / Q000187:drug effects; D000199:Actins / Q000502:physiology; D000205:Actomyosin / Q000502:physiology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure*; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004640:Emetine / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D008928:Mitochondria / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D000199:Actins; D014409:Tumor Necrosis Factor-alpha; D045304:Cytochromes c; D000205:Actomyosin; D004640:Emetine