The c-Jun n-terminal kinase/stress-activated protein kinase (JNK/SAPK) pathway in paclitaxel-induced apoptosis of cancer cells

The essential cellular functions associated with microtubules have led to a wide use of microtubule-interfering agents in cancer chemotherapy with promising results. Although the most well studied effect of microtubule-interfering agents is an arrest of cells at the G₂/M phase of the cell cycle,other effects may also exist. I have observed that paclitaxel (Taxol), docetaxel (Taxotere), vinblastine,vincristine, nocodazole and colchicine activate the c-Jun N-terminal kinase/stress-activated protein kinase(JNK/SAPK) signaling pathway in a variety of human cells. Activation of JNK/SAPK by microtubule interfering agents is dose-dependent and time-dependent and requires interactions with microtubules.Functional activation of the JNKK/SEKl-JNK/SAPK-c Jun cascade was demonstrated by cotransfection with a TPA-response element reporter construct and dominant negative (dn) signal transducers followed by chloramphenicol acetyl-transferase assays. Microtubule-interfering agents also activate both Ras and apoptosis signal-regulating kinase (ASKl), and coexpression of dn Ras and dn ASKl exerted individual and additive inhibition of JNK/SAPK activation by microtubule-interfering agents. These findings suggest that multiple signal transduction pathways are involved with cellular detection of microtubular disarray and subsequent activation of JNK/SAPK.To further examine the role of JNK/SAPK signaling cascades in apoptosis resulting from microtubule dysfunction induced by paclitaxel, I have coexpressed dn signaling proteins of theJNK/SAPK pathway (Ras, ASKl, Rac, JNKK, JNK) in human ovarian cancer cells with a selectable marker to analyze the apoptotic characteristics of cells expressing dn-vectors following exposure to paclitaxel. Expression of these dn signaling proteins had no effect on Bcl-2 phosphorylation, yet inhibited apoptotic changes induced by paclitaxel up to 16 h after treatment. Coexpression of these dna-signaling proteins had no protective effect after 48 h of paclitaxel treatment. These data indicate that: (i) activatedJNK/SAPK acts upstream of membrane changes and caspase-3 activation in paclitaxel-initiated apoptotic pathways, independently of cell cycle stage, (ii) activated JNK/SAPK is not responsible for paclitaxelinducedphosphorylation of Bcl-2, and (iii) apoptosis resulting from microtubule damage may comprise multiple mechanisms, including a JNK/SAPK-dependent early phase and a JNK/SAPK-independent latephase.