title: PU.1 Suppresses Th2 Cytokine Expression via Silencing of GATA3 Transcription in Dendritic Cells.
abstract: The transcription factor PU.1 is predominantly expressed in dendritic cells (DCs) and is essential for DC differentiation. Although there are several reports that PU.1 positively regulates the expression of DC-specific genes, whether PU.1 also has a suppressive effect on DCs is largely unknown. Here we demonstrate that PU.1 suppresses the expression of Th2 cytokines including IL-13 and IL-5 in bone marrow-derived DCs (BMDCs), through repression of the expression of GATA3, which is a master regulator of Th2 differentiations. When PU.1 siRNA was introduced into BMDCs, LPS-induced expression of IL-13 and IL-5 was increased along with upregulation of the constitutive expression of GATA2 and GATA3. The additional introduction of GATA3 siRNA but not of GATA2 siRNA abrogated PU.1 siRNA-mediated upregulation of IL-13 and IL-5. A chromatin immunoprecipitation assay showed that PU.1 bound to Gata3 proximal promoter region, which is more dominant than the distal promoter in driving GATA3 transcription in DCs. The degree of histone acetylation at the Gata3 promoter was decreased in PU.1 siRNA-introduced DCs, suggesting the involvement of PU.1 in chromatin modification of the Gata3 promoter. Treatment with a histone deacetylase (HDAC) inhibitor, trichostatin A, increased the degree of histone H3 acetylation at the Gata3 promoter and induced the subsequent expression of GATA3. Experiments using HDAC inhibitors and siRNAs showed that HDAC3 suppressed GATA3 expression. The recruitment of HDAC3 to the Gata3 promoter was decreased by PU.1 knockdown. LPS-induced IL-13 expression was dramatically reduced in BMDCs generated from mice lacking the conserved GATA3 response element, termed CGRE, which is an essential site for the binding of GATA3 on the Il-13 promoter. The degree of H3K4me3 at CGRE was significantly increased in PU.1 siRNA-transfected stimulated DCs. Our results indicate that PU.1 plays pivotal roles in DC development and function, serving not only as a transcriptional activator but also as a repressor.
authors.affiliation: Laboratory of Molecular Biology and Immunology, Department of Biological Science and Technology, Faculty of Industrial Science and Technology, Tokyo University of Science, 6-3-1 Niijuku, Katsushika-ku, Tokyo, 125-8585, Japan; Atopy Research Center, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Yashiro
authors.affiliation: Laboratory for Cytokine Regulation, Integrative Medical Science (IMS), RIKEN Yokohama Institute, Kanagawa, Japan; Division of Molecular Pathology, Research Institute for Biomedical Science, Tokyo University of Science, 2669 Yamazaki, Noda, Chiba, 278-0022, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Kubo
authors.affiliation: Atopy Research Center, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan.
authors.forename: Hideoki
authors.initials: H
authors.lastname: Ogawa
authors.affiliation: Atopy Research Center, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan.
authors.forename: Ko
authors.initials: K
authors.lastname: Okumura
authors.affiliation: Laboratory of Molecular Biology and Immunology, Department of Biological Science and Technology, Faculty of Industrial Science and Technology, Tokyo University of Science, 6-3-1 Niijuku, Katsushika-ku, Tokyo, 125-8585, Japan; Atopy Research Center, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan.
authors.forename: Chiharu
authors.initials: C
authors.lastname: Nishiyama
mesh_terms: D000107:Acetylation; D020816:Amino Acid Motifs; D000818:Animals; D016207:Cytokines / Q000235:genetics / Q000378:metabolism*; D003713:Dendritic Cells / Q000378:metabolism*; D005091:Exons; D050982:GATA1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050989:GATA2 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050990:GATA3 Transcription Factor / Q000235:genetics*; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D006655:Histone Deacetylases / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D018793:Interleukin-13 / Q000235:genetics / Q000378:metabolism; D015848:Interleukin-5 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D059372:Nucleotide Motifs; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D018418:Th2 Cells / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D000099232:Proto-Oncogene Protein Spi-1
chemical_list: D016207:Cytokines; D050982:GATA1 Transcription Factor; D050989:GATA2 Transcription Factor; D050990:GATA3 Transcription Factor; D006655:Histone Deacetylases; D006657:Histones; D018793:Interleukin-13; D015848:Interleukin-5; D011518:Proto-Oncogene Proteins; D034741:RNA, Small Interfering; D015534:Trans-Activators; D000099232:Proto-Oncogene Protein Spi-1