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Previous Article | Next Article 
Molecular and Cellular Biology, July 2004, p. 6040-6048, Vol. 24, No. 13
0270-7306/04/$08.00+0 DOI: 10.1128/MCB.24.13.6040-6048.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.
B Kinase Is an Essential Component of the Tpl2 Signaling Pathway
Department of Microbiology and Immunology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033
Received 21 October 2003/ Returned for modification 12 December 2003/ Accepted 8 April 2004
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ABSTRACT |
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B kinase (IKK), a key regulator of immune and inflammatory responses, is known as an effector kinase mediating activation of the transcription factor NF-B. Whether IKK also participates in other signaling events is not known. Here we show that IKK serves as an essential component of a signaling pathway that involves activation of the Tpl2 kinase and its downstream targets, MEK1 and ERK. Inhibition of IKKß in macrophages eliminates Tpl2 activation and ERK phosphorylation induced by lipopolysaccharide and tumor necrosis factor alpha. Using IKK-deficient murine fibroblasts, we further demonstrate that IKKß, but not IKK
, is required for Tpl2 activation. Moreover, this novel function of IKKß appears to involve phosphorylation and degradation of the Tpl2 inhibitor NF-B1/p105. These findings suggest that IKKß exerts its immune-regulatory functions by targeting different downstream signaling pathways. |
INTRODUCTION |
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B kinase (IKK) and three classes of mitogen-activated protein kinases (MAPK): the extracellular signal-regulated kinase (ERK), the c-jun N-terminal kinase (JNK), and p38 (36). These effector kinases each target the activation of specific transcription factors, which in turn cooperate in the activation of a large array of proinflammatory genes, such as those encoding the cytokines tumor necrosis factor (TNF-) and interleukin-1ß, the prostaglandin-generating enzyme cycloxygenase 2 (COX-2), and intracellular nitric oxide synthase. Binding of the inflammatory cytokines to their receptors on macrophages and other cell types can further amplify the innate immune response.
IKK is known as the effector kinase mediating activation of NF-B, a family of transcription factors that regulates macrophage activation as well as many other parts of the immune response (20, 34). The NF-B proteins are normally sequestered in the cytoplasm as inactive complexes by physical interaction with specific inhibitors, including IB and related proteins (3). Upon activation by cellular stimuli, IKK phosphorylates IBs, targeting these inhibitors for ubiquitination and proteasomal degradation, which allows released NF-B factors to move to the nucleus and exert their transactivation function (19). In mammals, the NF-B family consists of five members, RelA, RelB, c-Rel, NF-B1, and NF-B2, which function as homo- and heterodimers (33). A characteristic of NF-B1 and NF-B2 is that they are translated as large precursor proteins, p105 and p100, which upon proteasome-mediated processing generate the mature NF-B subunits p50 and p52, respectively (11, 33). In addition to serving as precursor proteins, p105 and p100 also function as IB molecules (26, 29). Recent studies suggest that the inducible processing of p100 controls a noncanonical NF-B signaling pathway involved in B-cell maturation and lymphoid organogenesis (31, 41). In contrast to the processing of p100, the processing of p105 is constitutive and occurs cotranslationally (24). Interestingly, a large proportion of p105 remains as the unprocessed form, which undergoes degradation in response to certain cellular signals (4, 13, 15, 16, 28). However, the physiological role of p105 degradation has not been well defined.
IKK was originally isolated as a holoenzyme complex composed of two catalytic subunits, IKK and IKKß, and one regulatory subunit, IKK
. Emerging evidence suggests that the different IKK components may also function in different kinase complexes (7, 40) and exert distinct functions (12). IKK plays an essential role in mediating p100 processing and, thus, the noncanonical pathway of NF-B activation (31). On the other hand, IKKß and IKK are required for the canonical NF-B signaling, specified by rapid degradation of IB and nuclear translocation of RelA-containing NF-B dimers (12). Genetic studies reveal that inactivation of IKKß, but not that of IKK, severely cripples the activation of NF-B by proinflammatory stimuli (22, 23, 37). IKKß has consistently been shown to function as a key regulator of inflammatory responses (6). It remains unknown, however, whether IKKß acts solely through NF-B or possesses additional targets.
Recent studies on MAPKs have identified an ERK signaling pathway that specifically responds to immune stimuli. A key regulatory component of this immunoresponsive pathway is a MAPK kinase kinase (MAP3K), Tpl2 (also named Cot) (8). Unlike the MAP3K Raf-1 that regulates the classical ERK pathway, Tpl2 does not respond to mitogens (39) but acts as a target of various immune stimuli, including the bacterial component LPS, the proinflammatory cytokine TNF-, and the B-cell stimulator CD40 ligand (8-10, 39). Genetic evidence suggests that the Tpl2/ERK signaling pathway plays a pivotal role in regulating the production of proinflammatory mediators by macrophages and antibody isotype switching in B cells (8-10, 39). However, how this novel signaling pathway is regulated has not been elucidated. Here we provide biochemical and genetic evidence that IKKß is a key upstream regulator of the Tpl2 signaling pathway. Using both pharmacological inhibitors and IKK-deficient cells, we demonstrate that IKKß as well as its regulatory subunit IKK are required for Tpl2 activation. These findings suggest that IKKß exerts its immune-regulatory function by targeting different downstream signaling pathways, including those leading to activation of NF-B and MAPK.
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MATERIALS AND METHODS |
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b1 gene (32) and their closely related wild-type control (B6 129PF2/J) were purchased from Jackson Laboratories. All experiments were done in accordance with Institutional Guidelines of the Pennsylvania State University College of Medicine.
Plasmid constructs, antibodies, and other reagents.
The retroviral expression vector encoding wild-type p105 was generated by cloning human p105 cDNA into the pCLXSN vector (27). PCLXSN-p105SSS/AAA was generated by site-directed mutagenesis to replace three serines (921, 923, and 927) located at the IKK phosphorylation site of p105 with alanines. The plasmid vectors encoding Tpl2 and IKKß were generated by cloning the corresponding cDNA into the pcDNA-hemagglutinin (HA) vector (14), and the bacterial expression vector encoding glutathione S-transferase (GST)-MEK1 was constructed by cloning human MEK1 into the pGEX4T-3 vector (Amersham/Pharmacia Biotech). GST-p105C and GST-p105C SSS/AAA were generated by cloning the C-terminal 535 amino acids of wild-type p105 or p105SSS/AAA into pGEX4T-1 vector (Amersham/Pharmacia Biotech). GST-IB(1-54) has been described previously (38). GST fusion proteins were produced in bacteria and were purified with glutathione-conjugated Sepharose beads (Amersham/Pharmacia Biotech) following the manufacturer's instruction.
The anti-p105 antibody was provided by Nancy Rice. The sources of the other antibodies were as described previously (39). The IKKß inhibitor PS1145 was kindly provided by Millennium Pharmaceuticals, Inc. (5), and the IKK inhibitor 15dPGJ2 was purchased from Biomol Research Company, Inc. Both inhibitors were dissolved in dimethyl sulfoxide (DMSO). LPS derived from Escherichia coli O127:B8 and phorbol myristate acetate (PMA) were obtained from Sigma, and murine TNF- was from BioSource. The recombinant IKK and IKKß, produced in insect cells, were kindly provided by Michael Karin.
ELISA.
Bone marrow-derived macrophages (BMDM) were seeded at a density of 106 cells/ml. After overnight starvation, cells were stimulated with LPS for 6 h in the presence or absence of P1145. Supernatant was then collected from the cells after the 6-h LPS stimulation and TNF- was measured by using an enzyme-linked immunosorbent assay (ELISA) kit from Pharmigen (OptEIA TNF- kit) following the manufacturer's protocol.
In vitro translation. The pcDNA-based Tpl2 and IKKß plasmid vectors were used for in vitro translations, because they carry the T7 promoter. The coupled TNT in vitro translation kit (Promega) was used.
Cells, in vitro stimulation, and retroviral infection.
BMDM were prepared and cultured as previously described (39). MEFs derived from wild-type and IKK-deficient mice were kindly provided by Michael Karin. The murine RAW264 macrophage cell line was from American Type Culture Collection. The cells were starved overnight in medium supplemented with 0.5% fetal bovine serum and then were stimulated for the indicated times. Following stimulation the cells were rapidly scraped into phosphate-buffered saline from culturing dishes, collected by centrifugation, and immediately preseeded for extract preparation. Retroviral infection of the nfb1–/– BMDM was as described previously (39).
Immunoblotting, coIP, and protein kinase assays. The procedures for immunoblotting, coimmunoprecipitation (coIP), and kinase assays were as previously described (39).
EMSA. Nuclear extracts were prepared from BMDM and were subjected to electrophoresis mobility shift assay (EMSA) with a 32P-radiolabeled B oligonucleotide as previously reported (35).
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RESULTS |
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To investigate the role of IKK in regulating inflammatory responses, we examined the effect of a selective IKK inhibitor, PS1145 (5, 17), on NF-B activation and production of inflammatory mediators in BMDM. As expected, both LPS and TNF- induced degradation of the prototypical NF-B inhibitor IB, with LPS being a stronger inducer (Fig. 1A, lanes 2 and 5). Preincubation of the cells with PS1145 partially inhibited the inducible degradation of IB by both LPS (lanes 3 and 4) and TNF- (lanes 6 and 7). Furthermore, EMSA revealed a similar inhibitory effect of PS1145 on the activation of NF-B (Fig. 1B). Using purified recombinant IKK components, we found that PS1145 selectively inhibited the kinase activity of IKKß but had no effect on the function of IKK (Fig. 1C). The inability of PS1145 to inhibit IKK may explain the incomplete block of NF-B activation in the drug-treated cells (Fig. 1B). We then examined the effect of PS1145 on two major proinflammatory mediators produced by LPS-stimulated macrophages, the cytokine TNF- and the prostaglandin-generating enzyme COX-2. In the absence of PS1145, LPS potently stimulated the secretion of TNF- (Fig. 1D) and expression of Cox-2 (Fig. 1E) by macrophages. More importantly, both of these LPS-mediated intracellular events were efficiently blocked by PS1145 (Fig. 1D and E).
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-stimulated ERK1 and ERK2 activation (Fig. 2B, lanes 1 to 3). Interestingly, however, this IKK inhibitor had no effect on ERK activation stimulated by the mitogen PMA (Fig. 2B, lanes 4 to 6). These results suggest that PS1145 inhibits the activation of ERK by specific inducers, namely the inflammatory elicitors LPS and TNF-.
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B (Fig. 2C) and activation of NF-B (data not shown) in LPS-treated cells. More importantly, inhibition of IKK by 15dPGJ2 resulted in the elimination of LPS-stimulated ERK1 and ERK2 activation (Fig. 2C). Similar to that of PS1145, the ERK-inhibitory function of 15dPGJ2 was specific, because it had little effect on LPS-stimulated activation of JNK and p38 (Fig. 2C) and PMA-stimulated ERK1 and ERK2 activation (data not shown). Together with the previous finding that the ERK pathway is essential for LPS-stimulated production of TNF- and COX-2 (8, 9), these results suggest that the potent anti-inflammatory effect of IKK inhibitors involves inhibition of both the NF-B and ERK signaling pathways. These findings also imply that IKK may function as an essential component of the LPS-responsive ERK signaling pathway. IKK inhibitors block activation of MAP3K Tpl2. The finding that IKK is dispensable for PMA-stimulated ERK activation suggests that IKK does not directly regulate ERK but may target an upstream component in the ERK signaling pathway. In this regard, ERK activation is known to be mediated by a MAP2K, MEK1, which in turn is regulated by the MAP3K Tpl2 in the response to inflammatory signals (8, 10, 39). We first investigated the role of IKK in mediating the activation of MEK1. Stimulation of macrophages by LPS resulted in potent activation of MEK1, as demonstrated by its in vivo phosphorylation (Fig. 3A). More importantly, this signaling event was completely blocked by the IKKß inhibitor PS1145 (Fig. 3A, lane 3). Similar to its effect on ERK activation, PS1145 did not inhibit the activation of MEK1 stimulated by PMA (Fig. 3A, lane 5). This result suggests that a more upstream kinase, instead of MEK1, may serve as the direct target of IKK.
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(10). We thus investigated whether the TNF--stimulated Tpl2 activation was also sensitive to IKK inhibitors. Like LPS, TNF- stimulated the kinase activity of Tpl2, as demonstrated by the in vitro (Fig. 3C, top panel) and in vivo (second panel) phosphorylation of MEK1 (lane 2). Although TNF- was a weaker Tpl2 inducer than LPS (data not shown), the TNF--stimulated Tpl2 activation was efficiently blocked by the IKK inhibitors (Fig. 3C, lane 3). The above-described studies performed with IKK inhibitors suggested that the activation of Tpl2 is under the control of IKK. However, we also considered the possibility that the IKK inhibitors might directly affect the catalytic activity of Tpl2. To exclude this possibility, we performed detailed titration studies to examine the inhibitory effect of PS1145 on the catalytic activity of IKKß and Tpl2. As expected, PS1145 efficiently inhibited the kinase activity of recombinant IKKß in vitro (Fig. 3D, top panel). In contrast, the catalytic activity of recombinant Tpl2 was not significantly affected by PS1145 at doses up to 15 µM (middle panel). Similarly, PS1145 did not inhibit MEK1-mediated phosphorylation of ERK (lower panel). Parallel studies using 15dPGJ2 similarly revealed that this IKK inhibitor had no inhibitory effect on Tpl2 or MEK1 (data not shown). Therefore, it is likely that the IKK inhibitors have no direct effect on the catalytic activity of Tpl2 or its downstream kinase MEK1 but rather block the activation of these kinases through inhibition of IKK. These results further suggested that IKK is required for activation of the Tpl2 signaling pathway.
Activation of Tpl2 and phosphorylation of ERK are preceded by IKK activation. If IKK is indeed required for Tpl2 activation, its activation should precede that of Tpl2. We performed kinetic studies to examine this possibility. Upon LPS stimulation, IKK was activated as early as 2.5 min, activity reached the maximal level around 5 min, and the activity persisted until at least 30 min poststimulation (Fig. 4, top panel, lanes 2 to 6). The activation of Tpl2 was delayed compared to that of IKK. The Tpl2 activity was first detected at 5 min after LPS stimulation and reached the maximal level around 15 min (middle panel, lanes 2 to 6). Consistent with the role of Tpl2 in MEK1 activation (8), in vivo phosphorylation of the MEK1 substrate, ERK1 and ERK2, was also more delayed than the activation of IKK (bottom panel, lanes 2 to 6).
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revealed that the overall signaling kinetics induced by this proinflammatory cytokine are more rapid and transient than that stimulated by LPS. Maximal IKK activation was detected at 2.5 min after TNF- stimulation, and the activity became largely diminished around 15 min (Fig. 4, top panel, lanes 8 to 12). Importantly, the activation of Tpl2 and phosphorylation of ERK again exhibited delayed kinetics (middle and lower panels, lanes 8 to 12).
Genetic evidence for the essential role of IKKß and IKK in Tpl2 activation.
To obtain genetic evidence for the regulatory role of IKK in Tpl2 activation, we examined this signaling event by using mouse embryonic fibroblasts (MEF) harboring deficiencies in different IKK subunits. Although the wild-type MEFs did not appreciably respond to LPS (data not shown), they supported the Tpl2 activation by TNF- (Fig. 5, upper panel, lanes 2 and 3). The MEFs deficient in IKK (IKK–/–) or IKKß (IKKß–/–) exhibited a residual basal activity of Tpl2 (lanes 4 and 7). Interestingly, the IKK–/– MEFs were fully responsive to TNF--induced Tpl2 activation (lanes 4 to 6), suggesting that this IKK component is dispensable for this signaling event. In contrast, the TNF--stimulated Tpl2 activation was completely defective in the IKKß–/– MEFs (lanes 7 to 9). Consistent with its essential role in modulating IKKß activation, the regulatory subunit IKK was also required for TNF--mediated Tpl2 activation (lanes 10 to 12). We also tried to analyze the phosphorylation of ERK in the mutant MEFs. Unfortunately, these MEFs exhibited high constitutive ERK activity even after serum starvation (data not shown). Nevertheless, such ERK activity is independent of Tpl2, because Tpl2 remains inducible in these cells (Fig. 5) and because the constitutive ERK activity is insensitive to the IKK inhibitor (data not shown). Nevertheless, these genetic studies clearly demonstrate that IKKß and IKK are required for Tpl2 activation.
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b1 gene product p105 forms a stable complex with Tpl2 (4) and functions as a physiological inhibitor of Tpl2 (39). The activation of Tpl2 appears to involve the release of its long isoform (Tpl2L) from p105, although the underlying mechanisms have not been resolved (39). We examined whether the IKK inhibitors affected this critical step of Tpl2 activation. Consistent with the results of previous studies (4, 39), both isoforms of Tpl2 (Tpl2L and Tpl2S) were coprecipitated with p105 in a coIP assay (Fig. 6A, second panel, lane 1), suggesting their stable association with p105. Further, in response to LPS stimulation, Tpl2L was rapidly released from the p105 complex (lanes 2 and 3). Remarkably, the dissociation of Tpl2 from p105 was completely blocked by both of the IKK inhibitors, PS1145 (Fig. 6A, second panel, lane 4) and 15dPGJ2 (data not shown).
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stimulation of wild-type MEFs resulted in efficient release of Tpl2L from p105 (Fig. 6B, second panel, lane 2). This step of Tpl2 signaling does not require IKK, because it remained normal in IKK–/– MEFs (lane 4). In contrast, the TNF--induced Tpl2 and p105 dissociation was completely blocked in the IKKß–/– (lane 6) and IKK–/– MEFs (lane 8). These genetic studies further confirmed the essential role for IKKß as well as the regulatory subunit (IKK) of canonical IKK in regulating the activation of Tpl2. Proteasome inhibitors block LPS-stimulated Tpl2/ERK signaling. We noticed that the LPS-stimulated release of Tpl2L from p105 was associated with a partial loss of p105 (Fig. 6A, top panel, lanes 2 and 3). Interestingly, the p105 degradation was blocked by the IKKß inhibitor PS1145 (lane 4). These findings suggest that the LPS-induced Tpl2 liberation and activation may result from IKK-mediated degradation of p105. We further examined this possibility by performing a more detailed time course experiment to correlate p105 degradation with Tpl2 activation, Tpl2L liberation, and degradation. A partial loss of p105 was noticeable at 10 min of LPS stimulation and became more prominent at 25 min (Fig. 7A, second panel). This dynamic change was correlated with the release of Tpl2L from p105 (third panel) and the activation of Tpl2 kinase activity (top panel). The kinetics of Tpl2L degradation (bottom panel) was evidently delayed compared to that of its liberation (third panel). These results support the hypothesis that activation of Tpl2 involves degradation of p105 and liberation of Tpl2L. We noticed that p105 was not completely degraded even after extended LPS stimulation (50 min), indicating that the degradation signal might preferentially target a proportion of p105. This idea is supported by the finding that the p105/Tpl2S complex was not significantly dissociated following LPS stimulation (Fig. 7A, third panel).
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-treated cells, although the activation of IKK was more transient (lanes 4 and 5). Further, the phosphorylation of p105 requires its C-terminal serines, because a mutant form lacking these serines (p105SSS/AAA) was only weakly phosphorylated (lane 7). To address the physiological role of p105 phosphorylation in Tpl2 activation, we employed a genetic model system involving reconstitution of the nfb1–/– macrophages with p105 or p105SSS/AAA. In this regard, it was previously shown that the nfb1–/– cells have a deficiency in Tpl2 expression and LPS-stimulated ERK activation, which can be rescued by expression of p105 (39). As expected, the steady expression of Tpl2 in nfb1–/– cells was restored upon p105 expression (Fig. 9B, lower panel, lane 2). The same result was obtained by expression of p105SSS/AAA (lane 3), thus suggesting the dispensability of p105 phosphorylation in Tpl2 stabilization. However, unlike the wild-type p105, the p105SSS/AAA mutant was unable to rescue the signaling defect of nfb1–/– macrophages in LPS-stimulated ERK activation (Fig. 9C, middle panel). This functional deficiency of p105SSS/AAA was apparently due to its inability to respond to LPS signal for degradation. Indeed, the wild-type p105 (Fig. 9C, upper panel, lane 4) but not p105SSS/AAA (lane 6) was degraded by the LPS signal. These genetic studies further suggest an essential role for IKK-mediated p105 phosphorylation in LPS-stimulated ERK signaling.
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DISCUSSION |
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B and different classes of MAPKs. Compelling evidence suggests that IKK plays a pivotal role in macrophage activation as well as in many other parts of immune responses (20, 34). Up to now, IKK has been thought to function as a specific effector kinase mediating activation of NF-B. The results presented in the present study, however, suggest that this key immune regulator also acts through activating other signaling pathways.
Using both IKK inhibitors and IKK-deficient MEFs, we have shown that IKK is required for activation of Tpl2, a MAP3K that specifically mediates MEK and ERK activation by immune stimuli (8, 10, 39). This novel function of IKK appears to be mediated by its canonical complex. Indeed, IKKß, the key component of the canonical IKK (12), is essential for Tpl2 activation. Further, the regulatory subunit of the canonical IKK complex, IKK, is also indispensable for this signaling function. In contrast, the activation of Tpl2 does not require the noncanonical IKK component IKK. We have shown that inhibition of IKKß by the inhibitor PS1145 does not completely eliminate LPS-stimulated NF-B activation. This is likely due to the functional compensation by IKK, as suggested by IKKß knockout studies (21). Surprisingly, however, the IKKß inhibitor is effective in blocking the Tpl2/ERK signaling in macrophages. This result suggests that the role of IKKß in Tpl2 activation cannot be compensated for by IKK. This idea is further supported by the finding that IKKß–/– MEFs are completely defective in Tpl2 activation.
Although our data clearly demonstrate the requirement for IKKß in Tpl2 activation, the role of IKKß in mediating ERK activation is less clear. Because ERK activation can be mediated through either Tpl2-dependent or -independent mechanisms, the effect of IKKß on this downstream pathway may vary among cell types and cellular inducers. In this regard, previous Tpl2 knockout studies suggest that ERK activation by LPS and TNF- in macrophages is dependent on Tpl2 (8, 9). We have consistently shown that inhibition of Tpl2 by the IKKß inhibitor abolishes ERK activation by LPS and TNF-. On the other hand, a recent study revealed that conditional knockout of IKKß in hepatocytes has no effect on ERK activation in response to LPS administration (25). Because LPS likely induces hepatocyte effects through TNF-, this finding indicates that IKKß may not be required for TNF--induced ERK activation in hepatocytes. Because a major role of Tpl2 is the regulation of ERK signaling in macrophages (8), mutant mice with IKKß deficiency in macrophages will be important for understanding the in vivo role of the IKKß/Tpl2 signaling pathway in macrophage activation and inflammatory responses.
It was previously shown that Tpl2 activation involves the release of its long isoform, Tpl2L, from the NF-B1 precursor protein p105 (39). Our present results suggest that the Tpl2L liberation requires p105 phosphorylation by IKKß, because Tpl2L cannot be liberated from a p105 mutant lacking its IKK phosphorylation site. Further, both the release of Tpl2L from p105 and the activation of Tpl2 catalytic activity are blocked in cells lacking IKKß or treated with IKKß inhibitors. Several lines of evidence indicate that the phosphorylation-coupled degradation of p105 contributes to Tpl2L liberation. First, partial degradation of p105 is associated with Tpl2L release. Second, inhibition of p105 degradation by proteasome inhibitors blocks the release of Tpl2L and the activation of Tpl2. Third, the defect in Tpl2L/p105 dissociation in IKKß–/– and IKK–/– MEFs is also associated with the blockade in p105 degradation. On the other hand, our data do not exclude the possibility of the involvement of proteolytic events in other signaling steps of Tpl2 activation.
It is presently unclear how cellular signals specifically target the dissociation of p105/Tpl2L complex but not that of the p105/Tpl2S complex. One possibility is that the p105/Tpl2 dissociation requires modifications (e.g., phosphorylation) of both p105 and Tpl2. In this regard, Tpl2L contains a unique N-terminal region that is missing in Tpl2S. The modification of Tpl2L may also serve to trigger its catalytic activity. Given that overexpressed Tpl2 induces p105 degradation (4), it is possible that the activated Tpl2L participates in signal-induced degradation of p105. Future studies will determine the precise sequence requirement for signal-induced liberation of Tpl2L from p105. Additionally, it is also important to examine whether any upstream kinases other than IKKß are required for Tpl2 activation. Given the high specificity of the Tpl2 activation signals, it is conceivable that activation of this signaling pathway requires both IKKß and additional regulators. Notwithstanding these issues, our results have uncovered an essential function of IKKß in Tpl2 activation and suggest the idea that the immunoregulatory function of IKKß is beyond the activation of NF-B. It remains possible that IKK may target additional downstream pathways in other parts of the immune response.
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ACKNOWLEDGMENTS |
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This work was supported by research grant 1 R01 AI057555, awarded to S.-C.S. and M.Z., from the National Institutes of Health. M.W. and W.R. were supported by a predoctoral and postdoctoral training grant (5T32CA60395-10) from the National Institutes of Health.
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FOOTNOTES |
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