Abstract
Persistent cell shrinkage, called apoptotic volume decrease (AVD), is a pivotal event of apoptosis. Activation of the volume-sensitive outwardly rectifying Cl− channel (VSOR) is involved in the AVD induction. On the other hand, activation of the MAP kinase (MAPK) cascade is also known to play a critical role in apoptosis. In the present study, we investigated the relationship between the AVD induction and the stress-responsive MAPK cascade activation during the apoptosis process induced by staurosporine (STS) in HeLa cells. STS was found to induce AVD within 2–5 min and phosphorylation of c-Jun N-terminal kinase (JNK) and p38 MAPK after over 20–30 min. VSOR blockers suppressed not only STS-induced AVD but also phosphorylation of JNK and p38 as well as activation of caspase-3/7. Moreover, a p38 inhibitor, SB203580, and a JNK inhibitor, SP600125, failed to affect STS-induced AVD, whereas these compounds reduced STS-induced activation of caspase-3/7. Also, treatment with ASK1-specific siRNA suppressed STS-induced caspase-3/7 activation without affecting the AVD induction. Furthermore, sustained osmotic cell shrinkage per se was found to trigger phosphorylation of JNK and p38, caspase activation, and cell death. Thus, it is suggested that activation of p38 and JNK is a downstream event of AVD for the STS-induced apoptosis of HeLa cells.
Highlights
The apoptosis process includes whole-cell shrinkage, activation of cysteine proteases called caspases, chromatin condensation, genome DNA fragmentation, and apoptotic body formation
It is suggested that activation of p38 and Jun N-terminal kinase (JNK) is a downstream event of apoptotic volume decrease (AVD) for the STS-induced apoptosis of HeLa cells
Our results show that volume-sensitive outwardly rectifying Cl− channel (VSOR) blockers reduced AVD induction and phosphorylation of stress-responsive Mitogen-activated protein (MAP) kinase (MAPK) induced by STS, but the suppression of these MAPKs did not impede AVD
Summary
The apoptosis process includes whole-cell shrinkage, activation of cysteine proteases called caspases, chromatin condensation, genome DNA fragmentation, and apoptotic body formation. Mitogen-activated protein (MAP) kinases (MAPKs), which are the family of kinases transducing signals from the cell membrane to the nucleus in response to a wide range of stimuli including stress, are known to be involved in apoptotic cell death [12]. Phosphorylation of ASK1 is involved in apoptosis [13,14,15,16,17] by inducing activation of JNK and p38 MAPK [14,18]. We first attempted in identifying the MAPK signaling pathway of the STS-induced apoptosis in HeLa cells. Our results show that VSOR blockers reduced AVD induction and phosphorylation of stress-responsive MAPKs induced by STS, but the suppression of these MAPKs did not impede AVD. It is concluded that AVD or persistent cell shrinkage precedes activation of stress-responsive MAPK activation in HeLa cells undergoing STS-induced apoptosis
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