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Long-range intrachromosomal interactions in the T helper type 2 cytokine locus

Abstract

The T helper type 2 (TH2) locus control region is important in the regulation of the genes encoding the cytokines interleukins 4, 5 and 13. Using the chromosome conformation capture technique, we found that in T cells, natural killer cells, B cells and fibroblasts, the promoters for the genes encoding TH2 cytokines are located in close spatial proximity, forming an initial chromatin core configuration. In CD4+ T cells and natural killer cells, but not B cells and fibroblasts, the TH2 locus control region participates in this configuration. The transcription factors GATA3 and STAT6 are essential for the establishment and/or maintenance of these interactions. Intrachromosomal interactions in the TH2 cytokine locus may form the basis for the coordinated transcriptional regulation of cytokine-encoding genes by the TH2 locus control region.

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Figure 1: Chromosome conformation capture assay of the TH2 cytokine locus.
Figure 2: Poised chromatin conformation of the TH2 cytokine gene loci and the TH2 LCR in T cells and NK cells but not in fibroblasts.
Figure 3: The RHS7 site of the TH2 LCR interacts with other hypersensitive sites of the locus and the promoters of the cytokine-encoding genes in T cells but not in fibroblasts.
Figure 4: Absence of STAT6 leads to impairment of interactions between the TH2 LCR and the rest of the TH2 cytokine locus.
Figure 5: The Il13 promoter interacts with the Il4 promoter and several DNase I–hypersensitive sites on the TH2 cytokine locus.
Figure 6: Calcium mobilization in 3T3 fibroblasts expressing GATA3 gain a secondary chromatin conformation in the TH2 locus.

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Acknowledgements

We thank F. Manzo for assistance with manuscript preparation; C. Hughes for technical assistance; P.E. Fields, G.R. Lee, S.T. Kim and L. Jones for discussions; F. Grosveld and W. de Laat for protocols on chromosome conformation capture technique and for suggestions on improving the manuscript; and Wyeth Laboratories for the donation of IL-12. Experiments in this study were approved by the Yale University Institutional Animal Care and Use Committee. Supported with a Cancer Research Institute fellowship (C.S.) and Howard Hughes Medical Institute (R.A.F.).

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Correspondence to Richard A Flavell.

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Supplementary information

Supplementary Fig. 1

The Il5 cytokine gene promoter interacts with the Il4 promoter and DNase I hypersensitive sites located at the 3′ end of the Il4 gene, in T cells. (PDF 182 kb)

Supplementary Fig. 2

The Rad50 gene promoter does not participate in the higher order chromatin conformation of the TH2 cytokine genes and the TH2 LCR. (PDF 139 kb)

Supplementary Fig. 3

DNase hypersensitive sites at the 5′ and 3′ of the Il4 gene participate in the higher order chromatin organization of the TH2 cytokine locus. (PDF 207 kb)

Supplementary Fig. 4

Cytokine production of all the cell types used in 3C assay. (PDF 121 kb)

Supplementary Fig. 5

3C assay-primer sequences. (PDF 352 kb)

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Spilianakis, C., Flavell, R. Long-range intrachromosomal interactions in the T helper type 2 cytokine locus. Nat Immunol 5, 1017–1027 (2004). https://doi.org/10.1038/ni1115

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