Yinling Hu

JNK2 and IKKβ Are Required for Activating the Innate Response to Viral Infection

Viral infection or double-stranded (ds) RNA induce interferons (IFN) and other cytokines. Transcription factors mediating IFN induction are known, but the signaling pathways that regulate them are less clear. We now describe two such pathways. The first pathway leading to NF-kappaB depends on the dsRNA-responsive protein kinase (PKR), which in turn activates IKB kinase (IKK) through the IKKbeta subunit. The second viral-and dsRNA-responsive pathway is PKR independent and involves Jun kinase (JNK) activation leading to stimulation of AP-1. Both IKKbeta and JNK2 are essential for efficient induction of type I IFN and other cytokines in response to viral infection or dsRNA. This study establishes a general role for these kinases in activation of innate immune responses.

IKKα controls formation of the epidermis independently of NF-κB

The IKKα and IKKβ catalytic subunits of IκB kinase (IKK) share 51% amino-acid identity and similar biochemical activities: they both phosphorylate IκB proteins at serines that trigger their degradation. IKKα and IKKβ differ, however, in their physiological functions. IKKβ and the IKKγ/NEMO regulatory subunit are required for activating NF-κB by pro-inflammatory stimuli and preventing apoptosis induced by tumour necrosis factor-α (refs 5,6,7,8,9,10,11). IKKα is dispensable for these functions, but is essential for developing the epidermis and its derivatives. The mammalian epidermis is composed of the basal, spinous, granular and cornified layers. Only basal keratinocytes can proliferate and give rise to differentiated derivatives, which on full maturation undergo enucleation to generate the cornified layer. Curiously, keratinocyte-specific inhibition of NF-κB, as in Ikkα-/- mice, results in epidermal thickening but does not block terminal differentiation. It has been proposed that the epidermal defect in Ikkα-/- mice may be due to the failed activation of NF-κB. Here we show that the unique function of IKKα in control of keratinocyte differentiation is not exerted through its IκB kinase activity or through NF-κB. Instead, IKKα controls production of a soluble factor that induces keratinocyte differentiation.

JNK2 is required for efficient T-cell activation and apoptosis but not for normal lymphocyte development

BACKGROUND The Jun N-terminal kinase (JNK) signaling pathway has been implicated in cell proliferation and apoptosis, but its function seems to depend on the cell type and inducing signal. In T cells, JNK has been implicated in both antigen-induced activation and apoptosis. RESULTS We generated mice lacking the JNK2 isozymes. The mutant mice were healthy and fertile but defective in peripheral T-cell activation induced by antibody to the CD3 component of the T-cell receptor (TCR) complex - proliferation and production of interleukin-2 (IL-2), IL-4 and interferon-gamma (IFN-gamma) were reduced. The proliferation defect was restored by exogenous IL-2. B-cell activation was normal in the absence of JNK2. Activation-induced peripheral T-cell apoptosis was comparable between mutant and wild-type mice, but immature (CD4(+) CD8(+)) thymocytes lacking JNK2 were resistant to apoptosis induced by administration of anti-CD3 antibody in vivo. The lack of JNK2 also resulted in partial resistance of thymocytes to anti-CD3 antibody in vitro, but had little or no effect on apoptosis induced by anti-Fas antibody, dexamethasone or ultraviolet-C (UVC) radiation. CONCLUSIONS JNK2 is essential for efficient activation of peripheral T cells but not B cells. Peripheral T-cell activation is probably required indirectly for induction of thymocyte apoptosis resulting from administration of anti-CD3 antibody in vivo. JNK2 functions in a cell-type-specific and stimulus-dependent manner, being required for apoptosis of immature thymocytes induced by anti-CD3 antibody but not for apoptosis induced by anti-Fas antibody, UVC or dexamethasone. JNK2 is not required for activation-induced cell death of mature T cells.

Epithelial-intrinsic IKKα expression regulates group 3 innate lymphoid cell responses and antibacterial immunity

Expression of IKKα in intestinal epithelial cells promotes IL-22 production by group 3 innate lymphoid cells, and this axis is essential for defense against Citrobacter rodentium infection and to limit intestinal inflammation in response to DSS treatment.

IKKα controls ATG16L1 degradation to prevent ER stress during inflammation

Inhibition of the I&kgr;B kinase complex (IKK) has been implicated in the therapy of several chronic inflammatory diseases including inflammatory bowel diseases. In this study, using mice with an inactivatable IKK&agr; kinase (Ikk&agr;AA/AA), we show that loss of IKK&agr; function markedly impairs epithelial regeneration in a model of acute colitis. Mechanistically, this is caused by compromised secretion of cytoprotective IL-18 from IKK&agr;-mutant intestinal epithelial cells because of elevated caspase 12 activation during an enhanced unfolded protein response (UPR). Induction of the UPR is linked to decreased ATG16L1 stabilization in Ikk&agr;AA/AA mice. We demonstrate that both TNF-R and nucleotide-binding oligomerization domain stimulation promote ATG16L1 stabilization via IKK&agr;-dependent phosphorylation of ATG16L1 at Ser278. Thus, we propose IKK&agr; as a central mediator sensing both cytokine and microbial stimulation to suppress endoplasmic reticulum stress, thereby assuring antiinflammatory function during acute intestinal inflammation.

IKKα Promotes Intestinal Tumorigenesis by Limiting Recruitment of M1-like Polarized Myeloid Cells

The recruitment of immune cells into solid tumors is an essential prerequisite of tumor development. Depending on the prevailing polarization profile of these infiltrating leucocytes, tumorigenesis is either promoted or blocked. Here, we identify IκB kinase α (IKKα) as a central regulator of a tumoricidal microenvironment during intestinal carcinogenesis. Mice deficient in IKKα kinase activity are largely protected from intestinal tumor development that is dependent on the enhanced recruitment of interferon γ (IFNγ)-expressing M1-like myeloid cells. In IKKα mutant mice, M1-like polarization is not controlled in a cell-autonomous manner but, rather, depends on the interplay of both IKKα mutant tumor epithelia and immune cells. Because therapies aiming at the tumor microenvironment rather than directly at the mutated cancer cell may circumvent resistance development, we suggest IKKα as a promising target for colorectal cancer (CRC) therapy.

The AP-1 transcription factor JunB is essential for multiple myeloma cell proliferation and drug resistance in the bone marrow microenvironment

Despite therapeutic advances, multiple myeloma (MM) remains an incurable disease, predominantly because of the development of drug resistance. The activator protein-1 (AP-1) transcription factor family has been implicated in a multitude of physiologic processes and tumorigenesis; however, its role in MM is largely unknown. Here we demonstrate specific and rapid induction of the AP-1 family member JunB in MM cells when co-cultured with bone marrow stromal cells. Supporting a functional key role of JunB in MM pathogenesis, knockdown of JUNB significantly inhibited in vitro MM cell proliferation and survival. Consistently, induced silencing of JUNB markedly decreased tumor growth in a murine MM model of the microenvironment. Subsequent gene expression profiling revealed a role for genes associated with apoptosis, DNA replication and metabolism in driving the JunB-mediated phenotype in MM cells. Importantly, knockdown of JUNB restored the response to dexamethasone in dexamethasone-resistant MM cells. Moreover, 4-hydroxytamoxifen-induced activation of a JunB-ER fusion protein protected dexamethasone-sensitive MM cells against dexamethasone- and bortezomib-induced cytotoxicity. In summary, our results demonstrate for the first time a specific role for AP-1/JunB in MM cell proliferation, survival and drug resistance, thereby strongly supporting that this transcription factor is a promising new therapeutic target in MM.

Chronic intestinal inflammation in mice expressing viral Flip in epithelial cells

Viruses are present in the intestinal microflora and are currently discussed as a potential causative mechanism for the development of inflammatory bowel disease. A number of viruses, such as Human Herpesvirus-8, express homologs to cellular FLIPs, which are major contributors for the regulation of epithelial cell death. In this study we analyzed the consequences of constitutive expression of HHV8-viral FLIP in intestinal epithelial cells (IECs) in mice. Surprisingly, expression of vFlip disrupts tissue homeostasis and induces severe intestinal inflammation. Moreover vFlipIEC-tg mice showed reduced Paneth cell numbers, associated with excessive necrotic cell death. On a molecular level vFlip expression altered classical and alternative NFκB activation. Blocking of alternative NFκB signaling by deletion of Ikka in vivo largely protected mice from inflammation and Paneth cell loss induced by vFLIP. Collectively, our data provide functional evidence that expression of a single viral protein in IECs can be sufficient to disrupt epithelial homeostasis and to initiate chronic intestinal inflammation.