Cutaneous malignant melanoma is a highly aggressive and frequently chemoresistant cancer, whose incidence continues to rise. Epidemiological studies reveal that the major etiological melanoma risk factor is ultraviolet (UV) solar radiation, with the highest risk associated with intermittent burning doses, especially during childhood1,2. We have experimentally validated these epidemiological findings using the hepatocyte growth factor/scatter factor (HGF/SF) transgenic mouse model, which develops lesions in stages highly reminiscent of human melanoma with respect to biological, genetic and etiologic criteria, but only when irradiated as neonatal pups with UVB, not UVA3,4. However, mechanisms underlying UVB-initiated, neonatal-specific melanomagenesis remain largely unknown. Here we introduce a mouse model permitting fluorescence-aided melanocyte imaging and isolation following in vivo UV irradiation. We use expression profiling to show that activated neonatal skin melanocytes isolated following a melanomagenic UVB dose bear a distinct, persistent interferon response signature, including genes associated with immunoevasion. UVB-induced melanocyte activation, characterized by aberrant growth and migration, was abolished by antibody-mediated systemic blockade of interferon-γ (IFN-γ), but not type-I interferons. IFN-γ was produced by macrophages recruited to neonatal skin by UVB-induced ligands to the chemokine receptor Ccr2. Admixed recruited skin macrophages enhanced transplanted melanoma growth by inhibiting apoptosis; notably, IFN-γ blockade abolished macrophage-enhanced melanoma growth and survival. IFN-γ-producing macrophages were also identified in 70% of human melanomas examined. Our data reveal an unanticipated role for IFN-γ in promoting melanocytic cell survival/immunoevasion, and suggest that IFN-γ-R signaling represents a novel therapeutic melanoma target.

We analyzed interleukin (IL) 12 and IL-23 production by monocyte-derived dendritic cells (mono-DCs). Mycobacterium tuberculosis H37Rv and zymosan preferentially induced IL-23. IL-23 but not IL-12 was efficiently induced by the combination of nucleotide-binding oligodimerization domain and Toll-like receptor (TLR) 2 ligands, which mimics activation by M. tuberculosis, or by the human dectin-1 ligand beta-glucan alone or in combination with TLR2 ligands, mimicking induction by zymosan. TLR2 ligands inhibited IL-12 and increased IL-23 production. DC priming with interferon (IFN) gamma strongly increased IL-12 production, but was not required for IL-23 production and inhibited IL-23 production induced by beta-glucan. The pattern of IL-12 and IL-23 induction was reflected in accumulation of the IL-12p35 and IL-23p19 transcripts, respectively, but not IL-12/23p40. Although IL-23, transforming growth factor beta, and IL-6 contained in the supernatants of activated mono-DCs played a role in the induction of IL-17 by human CD4(+) T cells, IL-1beta, in combination with one or more of those factors, was required for IL-17 production, and its production determined the differential ability of the stimuli used to elicit mono-DCs to produce soluble factors directing IL-17 production. Thus, the differential ability of pathogens to induce antigen-presenting cells to produce cytokines regulates the immune response to infection

NFAT proteins constitute a family of transcription factors involved in mediating signal transduction. Using a panel of specific antisera in immunoprecipitation assays, we found that NFATp (135 kDa) is constitutively expressed in normal human T cells, while synthesis of NFATc (predominant form of 86 kDa) is induced by ionomycin treatment. NFAT4/x was very weakly expressed in unstimulated cells, and its level did not increase upon treatment with activating agents. NFAT3 protein was not observed under any conditions. Highermolecular-weight species of NFATc (of 110 and 140 kDa) were also detected. In addition, translation of NFATc mRNA apparently initiates at two different AUG codons, giving rise to proteins that differ in size by 36 amino acids. Additional size heterogeneity of both NFATc and NFATp results from phosphorylation. In contrast to ionomycin treatment, exposure of cells to phorbol myristate acetate (PMA) plus anti-CD28 did not induce NFATc, indicating that under these conditions, interleukin-2 synthesis by these cells is apparently independent of NFATc. In DNA binding assays, both PMA plus anti-CD28 and PMA plus ionomycin resulted in nuclear NFAT. Surprisingly, the PMA-ionomycin-induced synthesis of NFATc that was detected by immunoprecipitation was not mirrored in the DNA binding assays: nearly all of the activity was due to NFATp. This is the first study of expression of all family members at the protein level in normal human T cells.NFAT (nuclear factor of activated T cells) is implicated in regulation of interleukin-2 (IL-2) gene transcription (for reviews, see references 13 and 30). In addition, NFAT-binding sites have been identified in the regulatory regions of various other cytokine genes, including the IL-4 (3, 33, 38), tumor necrosis factor alpha (7, 23), and IL-3/granulocyte-macrophage colony-stimulating factor (4, 22) genes. Though originally found in T cells, NFAT DNA-binding activity and/or protein has now been found in other cell types, including B cells (2,40,41,45), mast cells (29), natural killer (NK) cells (1), and a neuronal cell line and certain regions of the brain (8). Thus, NFAT is likely to play an important role in the regulation of a variety of genes in a number of different cell types.To date, cDNAs from four different NFAT genes (NFATp, NFATc, NFAT3, and NFAT4/NFATx) have been cloned, and they constitute a related but quite divergent family (9,11,18,21,24,26,27). The family in turn is weakly related to the Rel/NF-B family of transcription factors over a 300-aminoacid region called the Rel homology domain (RHD). NFAT sequences in this region govern DNA binding and association with the AP-1 transcription factor (12), and within the RHD, sequence conservation among the NFAT proteins is very high. Upstream of the RHD, NFAT proteins are less closely related, but they do share several serine-and proline-rich segments. Downstream of the RHD, NFAT proteins are variable in length and in sequence.The hallmark of NFAT activity is its inducibility by agents that increase intracellular Ca...

Summary Interleukin-12 (IL-12) and IL-23 share a common chain. Yet, their production in response to pathogens is differentially regulated, and their functions are distinct and often antithetic. IL-12 is involved in the induction or amplification of the T-helper type 1 (Th1) response, whereas IL-23 has been associated with the generation of the Th17 response and IL-17 production. Mycobacterium tuberculosis and yeast zymosan induce IL-23, but in absence of other stimuli, no IL-12 is induced in human dendritic cells (DCs). The stimulation of IL-23 by M. tuberculosis was mostly explained by the triggering of Toll-like receptor 2 (TLR2) and the cytoplasmic receptor nucleotide oligomerization domain-containing protein 2 (NOD2), whereas zymosan induces IL-23 primarily by stimulating the β-glucan receptor dectin-1 alone or in combination with TLR2. IL-23, IL-6, transforming growth factor-β (TGF-β), and IL-1β in supernatants from activated human DCs induce human naive CD4+ T cells to produce IL-17. These data are consistent with various recent reports that TGF-β is an inducer of IL-17 production both in human and mouse cells. However, IL-1 is necessary in combination with some or all of the other cytokines to induce IL-17 production in human T cells. The ability of various stimuli to induce Th17 cells depends not only on their induction of IL-23, IL-6, and TGF-β production in DCs but also on their ability to activate directly or indirectly the inflammasome and to induce IL-1β.