The proinflammatory cytokine tumour necrosis factor-alpha (TNF-alpha) regulates immune responses, inflammation and programmed cell death (apoptosis). The ultimate fate of a cell exposed to TNF-alpha is determined by signal integration between its different effectors, including IkappaB kinase (IKK), c-Jun N-terminal protein kinase (JNK) and caspases. Activation of caspases is required for apoptotic cell death, whereas IKK activation inhibits apoptosis through the transcription factor NF-kappaB, whose target genes include caspase inhibitors. JNK activates the transcription factor c-Jun/AP-1, as well as other targets. However, the role of JNK activation in apoptosis induced by TNF-alpha is less clear. It is unknown whether any crosstalk occurs between IKK and JNK, and, if so, how it affects TNF-alpha-induced apoptosis. We investigated this using murine embryonic fibroblasts that are deficient in either the IKKbeta catalytic subunit of the IKK complex or the RelA/p65 subunit of NF-kappaB. Here we show that in addition to inhibiting caspases, the IKK/NF-kappaB pathway negatively modulates TNF-alpha-mediated JNK activation, partly through NF-kappaB-induced X-chromosome-linked inhibitor of apoptosis (XIAP). This negative crosstalk, which is specific to TNF-alpha signalling and does not affect JNK activation by interleukin-1 (IL-1), contributes to inhibition of apoptosis.

JNK has been suggested to be proapoptotic, antiapoptotic, or have no role in apoptosis depending on the cell type and stimulus used. The precise mechanism of JNK action, under conditions when it promotes cell survival, is not entirely clear. Here, we report that JNK is required for IL-3-mediated cell survival through phosphorylation and inactivation of the proapoptotic Bcl-2 family protein BAD. IL-3 withdrawal-induced apoptosis is promoted by inhibition of JNK but suppressed by expression of a constitutively active JNK. JNK phosphorylates BAD at threonine 201, thereby inhibiting BAD association with the antiapoptotic molecule BCL-X(L). IL-3 induces BAD phosphorylation at threonine 201, and replacement of threonine 201 by alanine generates a BAD mutant, which promotes IL-3 withdrawal-induced apoptosis. Thus, our results provide a molecular mechanism by which JNK contributes to cell survival.

Two ubiquitously expressed isoforms of c-Jun N-terminal protein kinase (JNK), JNK1 and JNK2, have shared functions and different functions. However, the molecular mechanism is unknown. Here we report that JNK1, but not JNK2, is essential for tumor necrosis factor alpha (TNF-α)-induced c-Jun kinase activation, c-Jun expression, and apoptosis. Using mouse fibroblasts deficient in either Jnk1 or Jnk2, we found that JNK1 was activated by TNF-α, whereas JNK2 activation was negligible. In addition, JNK2 interfered with JNK1 activation via its “futile” phosphorylation by upstream kinases. Consequently, expression and activation of c-Jun, which depends on JNK activity, were impaired in Jnk1 null cells but enhanced in Jnk2 null cells. TNF-α-induced apoptosis was also suppressed in Jnk1 null fibroblasts but increased in Jnk2 null cells. Thus, our results provide a molecular mechanism underlying the different biological functions of JNK isoforms