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A transcription factor PU.1 is critical for Ccl22 gene expression in dendritic cells and macrophages.

ABSTRACT: The chemokine CCL22 is predominantly produced by dendritic cells (DCs) and macrophages. CCL22 acts on CCR4-expressing cells including Th2 and Treg. Although a correlation between the CCL22-CCR4 axis and allergic diseases has been established, the mechanism of monocyte lineage-specific Ccl22 gene expression is largely unknown. In the current study, we investigated transcriptional regulation of the Ccl22 gene in DCs and macrophages. Using reporter assays, we identified the critical cis-enhancing elements at 21/-18 and -10/-4 in the Ccl22 promoter. Electrophoretic mobility shift assays proved that transcription factor PU.1 directly binds to the cis-elements. Knockdown of PU.1 markedly decreased Ccl22 expression in bone marrow-derived DCs (BMDCs) and BM macrophages (BMDMs). Chromatin immunoprecipitation assays revealed that PU.1 bound to the Ccl22 promoter in not only BMDCs and BMDMs, but also splenic DCs and peritoneal macrophages. LPS stimulation increased the amount of PU.1 recruited to the promoter, accompanied by upregulation of the Ccl22 mRNA level, which was diminished by Spi1 knockdown. We identified similar cis-elements on the human CCL22 promoter, which were bound with PU.1 in human monocytes. Taken together, these findings indicate that PU.1 transactivates the Ccl22 gene in DCs and macrophages by directly binding to the two elements in the promoter.

SUBMITTER: Yashiro T

PROVIDER: S-EPMC6361964 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description: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.

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A transcription factor PU.1 is critical for Ccl22 gene expression in dendritic cells and macrophages.

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