JNK inhibitor SP600125 promotes the formation of polymerized tubulin, leading to G2/M phase arrest, endoreduplication, and delayed apoptosis

Abstract

The JNK inhibitor SP600125 strongly inhibits cell proliferation in many human cancer cells by blocking cell-cycle progression and inducing apoptosis. Despite extensive study, the mechanism by which SP600125 inhibits mitosis-related effects in human leukemia cells remains unclear. We investigated the effects of SP600125 on the inhibition of cell proliferation and the cell cycle, and on microtubule dynamics in vivo and in vitro. Treatment of synchronized leukemia cells with varying concentrations of SP600125 results in significant G 2 /M cell cycle arrest with elevated p21 levels, phosphorylation of histone H3 within 24 h, and endoreduplication with elevated Cdk2 protein levels after 48 h. SP600125 also induces significant abnormal microtubule dynamics in vivo. High concentrations of SP600125 (200 μM) were required to disorganize microtubule polymerization in vitro. Additionally, SP600125-induced delayed apoptosis and cell death was accompanied by significant poly ADP-ribose polymerase (PARP) cleavage and caspase-3 activity in the late phase (at 72 h). Endoreduplication showed a greater increase in ectopic Bcl-2-expressing U937 cells at 72 h than in wild-type U937 cells without delayed apoptosis. These results indicate that Bcl-2 suppresses apoptosis and SP600125-induced G 2 /M arrest and endoreduplication. Therefore, we suggest that SP600125 induces mitotic arrest by inducing abnormal spindle microtubule dynamics.

Msx2 mediates the inhibitory action of TNF-α on osteoblast differentiation

“…TNF-α treatment increased the phosphorylation of JNK and its downstream target, c-Jun, within five min ( Figure 5A). Although a previous report has shown that SP600125, a JNK inhibitor, promotes apoptosis in human leukemia cells (Moon et al, 2009), it did not exert cytotoxic effects on C2C12 cells at 10 μM concentration ( Figure 5B). RT-PCR and immunoblot analyses revealed that Msx2 induction by TNF-α was not affected by SP600125 treatment.…”

Dependency of 2-methoxyestradiol-induced mitochondrial apoptosis on mitotic spindle network impairment and prometaphase arrest in human Jurkat T cells

“…The monomeric fraction was prepared by extracting cells in monomeric extraction buffer (20 mM PIPES, 0.14 M NaCl, 1 mM MgCl 2 , 1 mM EGTA, 0.5% NP-40, and 0.5 mM PMSF, pH 6.8) as previously described [34] with some modifications. After centrifugation at 13,000 Â g for 10 min at room temperature, the NP-40-soluble fraction containing monomeric tubulin was collected.…”

Activation of c-Jun N-Terminal Kinase (JNK) during Mitosis in Retinal Progenitor Cells

“…Our data suggest that JNK may be required for the separation or localization of the centrosome in the cell poles, therefore controlling the separation of chromatids at the outset of anaphase, and possibly helping to prevent polyploidy. It has, indeed, been shown that inhibition of JNK results in polyploidy and apoptosis of cultured cell lines [37], [40]. Thus, JNK may be important to maintain genomic stability and prevent programmed cell death in progenitor cells during retinal development.…”

“…The data suggest that the JNK signaling pathway may be involved in controlling the M-phase of the cell cycle [35]–[40]. Most studies, however, were done in either tumor cell lines [20]–[22], [36]–[41] or dissociated cells [23], [24], [35], [42], and little is known of the role of this enzyme in cell proliferation in the central nervous system [43]–[45].…”