Innate immune cells such as macrophages sense the presence of microbial infection by pattern-recognition receptors (PRRs) and mount anti-microbial responses. The Toll-like receptor (TLR) family is one of the best-characterized PRR families recognizing various pathogens such as bacteria, viruses, protozoa and fungi. TLRs play a critical role in evoking innate immune responses to infection by various pathogens leading to production of inflammatory mediators, including proinflammatory cytokines, chemokines and interferons (IFNs). It is well known that macrophages are functionally polarized into M1 and M2 cells in response to infection with microorganisms and host mediators. M1 macrophages produce high levels of tumor necrosis factor (TNF) and nitric oxide (NO) by expressing inducible NO synthase (iNOS), and are critical for clearing bacterial, viral and fungal infections. Macrophages also play an important role in responses to parasite infection, tissue remodeling, angiogenesis and tumor progression. These macrophages are called alternatively activated macrophages or M2 macrophages. M2 macrophages are characterized by their high expression of Arginase-1 (Arg1), chitinase-like Ym1 (Chi3l3), found in inflammatory zone 1 (Fizz1 also known as Retnla), mannose receptor (MR, also known as CD206), and chemokines such as CCL17, CCD24,. The PRR system responsible for the recognition of helminth infection and M2 polarization has yet to be identified.
  Cytokines and growth factors have been implicated in the reprogramming of M1 and M2 macrophages. Whereas IFN-γ, produced by activated T cells and TLR ligands, induces M1 macrophage generation, stimulation of macrophages with IL-4 or IL-13 induces M2 type macrophages. In addition, immune complexes, IL-10 and glucocorticoid or secosteroid hormones are also known to generate M2 macrophages. Among growth factors, treatment of bone marrow (BM) cells with granulocyte macrophage-colony stimulation factor (GM-CSF) and macrophage-colony stimulation factor (M-CSF) leads to generation of M1 and M2 cells, respectively.
  TLRs trigger intracellular signaling pathways, inducing activation of a set of transcription factors, such as NF-κB, AP-1, C/EBPβ, PU.1 and IFN-regulatory factors (IRFs). These transcription factors cooperatively upregulate the expression of multiple genes such as proinflammatory cytokines, leading to M1 macrophage polarization. Proteins induced by TLR signaling are known to modulate inflammatory responses. IκBζ, a IκB family member, positively regulates certain genes such as IL-6 by interacting with NF-κBp50 or inducing histone modification, while ATF3 negatively regulates expression. Conversely, transcription factors such as STAT6 and peroxisome proliferator-activated receptor (PPAR) γ are involved in polarization of M2 macrophages.
  In addition to regulation by transcription factors themselves, epigenetic regulation is found to be essential for controlling proper gene expression. Histone modifications, particularly at the N-terminal tails, and dynamic chromatin remodeling have been shown to be important for controlling sets of genes. In the case of histone modification, trimethylation of histone 3 lysine 4 (H3K4) is associated with active gene transcription, whereas trimethylation of H3K9, H3K27 and H4K20 is linked to silencing of gene expression. The methylation of H3K27 is mediated by the Polycomb repressive complex 2 (PRC2) composed of Ezh2, Suz12 and Eed23. Proteins harboring a Jumonji-C (JmjC) domain, Jmjd3, UTX and UTY, are known to act as H3K27 demethylases catalyzing H3K27me3 (tri-methylated) to HeK27me1 (mono-methylated).
  It has been reported that the expression of Jmjd3 (also known as Kdm6b) is induced in macrophages by TLR stimuli in an NF-κB-dependent fashion. We also identified Jmjd3 as an early TLR-inducible gene in mouse macrophages by microarray analysis. H3K27 tri-methylation is implicated in the silencing of gene expression and it has been shown that Jmjd3 is recruited to transcription start sites (TSS) that are characterized by high levels of RNA polymerase II and H3K4me3. JMJD3 is reported to fine-tune macrophage activation by controlling Bmp2 and Hox expression. Furthermore, JMJD3 has been implicated in the control of development by the regulation of Hox, and oncogenesis by promoting the expression of Ink4a. Here we report the role of JMJD3 in vivo in controlling M2 macrophage polarization and identify Irf4 as a JMJD3 target gene critical for the regulation of macrophages.


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