The transcription factor NF-κB is a dimeric complex composed of members of the Rel family of proteins (12
). The classic form of NF-κB is a dimer consisting of the transcriptionally inactive p50 subunit and the p65/RelA (p65) subunit, which contains a potent, C-terminal transactivation domain (12
). NF-κB activity is suppressed primarily through interactions with various IκB proteins that promote the cytoplasmic localization of NF-κB (12
). The activation of NF-κB is achieved when signals, such as those elicited by tumor necrosis factor (TNF), activate a kinase known as IKK that causes the phosphorylation of IκB proteins (16
). This phosphorylation triggers the ubiquitination and subsequent degradation of the IκB proteins, allowing for the nuclear accumulation of NF-κB (12
). Gene-specific transcription by NF-κB is achieved through the recognition of distinct DNA binding sites in genes typically associated with inflammatory and immune responses as well as cell cycle regulation and apoptosis (2
Recently it was demonstrated that NF-κB is a potent regulator of apoptosis. Evidence from a number of groups has indicated that NF-κB suppresses apoptosis induced by TNF and other apoptotic stimuli by inhibiting the activation of the cell death caspase cascade as well as by inhibiting the release of cytochrome c
from the mitochondria (3
). The inhibition of apoptosis by NF-κB appears to be largely transcriptional, since several antiapoptotic genes, such as those encoding Bcl-xL, c-IAP1, c-IAP2, A1/Bfl-2, and the death domain proteins TRAF1 and TRAF2, have been shown to be transcriptionally regulated by NF-κB (7
). Therefore, when cells that lack functional NF-κB are treated with TNF, a significant proportion of cells undergo apoptosis (3
). Interestingly, cell death under these circumstances never reaches 100%, so it has been speculated that an additional pathway activated by TNF may play an antiapoptotic role when NF-κB is suppressed.
TNF is a key inflammatory cytokine that regulates signal transduction cascades, gene expression, and apoptosis. The biological effects of TNF are mediated through two distinct cell surface receptors, TNF-RI and TNF-RII (1
). Ligand-induced trimerization of the TNF receptors leads to the recruitment of the TNF-receptor-associated death domain protein, TRADD, which serves to recruit additional proteins involved in the activation of specific signal transduction pathways (1
). For example, when FADD is recruited to TRADD, a caspase cascade is initiated through the recruitment of caspase-8. In addition, binding of TRAF2 to TRADD leads to the recruitment of additional proteins that can activate both the stress-activated protein kinase cascade, which includes Jun kinase (JNK) (9
), and as the NF-κB signaling pathway (1
). Therefore, the stimulation of cells with TNF can have both apoptotic and antiapoptotic consequences and can lead to the simultaneous activation of different pathways. How these pathways interact and potentially regulate each other may have a profound effect on the activity of a particular signaling pathway as well as on the outcome of cell death.
Like NF-κB, JNK and its downstream target, c-Jun, have each been implicated in the control of cell death, although their roles have proven to be complex. In particular, JNK has been shown to either positively or negatively regulate cell death depending on the biological context. For instance, JNK-deficient (JNK1 and JNK2 null) embryonic fibroblasts are blocked in UV-induced apoptosis, while animals that are null for JNK exhibit enhanced forebrain cell apoptosis (17
). Also, JNK has been shown to be involved in the protection of cells against taxol-induced cell death by epidermal growth factor (22
). Importantly, c-Jun, the downstream effector of JNK, has also proven to play both pro- and antiapoptotic roles. c-Jun can prevent apoptosis during hepatogenesis, but it is necessary for excitotoxin-induced cell death in neurons (13
). Furthermore, depending on the differentiation state of PC12 cells, c-Jun can function in a positive or negative manner towards cell death (20
). It is these disparities that led us to ask whether the activation status of NF-κB might determine the pro- or antiapoptotic status of JNK or other members of the stress kinase signaling pathway.
We show, as other groups have, that TNF leads to the rapid and transient activation of JNK in wild-type cells. However, the transient nature of this activation depends on the p65 subunit of NF-κB, since JNK activity in response to TNF is sustained in cells that lack functional NF-κB. In addition, this regulation requires the transcription function of the p65 subunit and appears to involve the inhibition of upstream signals that control JNK activity but not of those that regulate another stress-activated kinase, p38. Therefore, based on the involvement of JNK in cell death, we speculated that the sustained activation of JNK in NF-κB null cells could play an antiapoptotic role, possibly explaining the lack of complete TNF-induced cell death in p65−/−
fibroblasts. In accordance, the use of inhibitors to suppress sustained JNK activity in cells that lack functional NF-κB results in enhanced cell death in response to TNF. Therefore, the JNK pathway appears to inhibit cell death when the antiapoptotic role for NF-κB is suppressed. Recently it has been shown that NF-κB negatively modulates JNK activity (15
). However, in contrast to our studies, those studies indicate that deregulated JNK activity, resulting from inhibition of NF-κB, provides a proapoptotic signal in response to TNF. Two of these papers (30
) indicate that the ability of NF-κB to regulate either XIAP or GADD45β suppresses JNK activity. However, a recent paper indicates that XIAP antiapoptotic function requires JNK1 activation (27
). Our data provide new evidence of functional cross-regulation between NF-κB and the JNK pathway at a level upstream of JNK-specific activation and provide a rationale to explain why loss of NF-κB does not fully sensitize cells to the apoptotic potential of TNF.