Critical Role of Transcription Factor PU.1 in the Function of the OX40L/TNFSF4 Promoter in Dendritic Cells.

Takuya Yashiro,Mutsuko Hara,Hideoki Ogawa,Chiharu Nishiyama,Ko Okumura

doi:10.1038/srep34825

Takuya Yashiro, Mutsuko Hara + Show 3 more

Open Access

https://doi.org/10.1038/srep34825

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Abstract

PU.1 is a hematopoietic lineage-specific transcription factor belonging to the Ets family. We investigated the role of PU.1 in the expression of OX40L in dendritic cells (DCs), because the regulatory mechanism of cell type-specific expression of OX40L, which is mainly restricted to antigen-presenting cells, is largely unknown despite the critical involvement in Th2 and Tfh development. PU.1 knockdown decreased the expression of OX40L in mouse DCs. Chromatin immunoprecipitation (ChIP) assays demonstrated that PU.1 constitutively bound to the proximal region of the OX40L promoter. Reporter assays and electrophoretic mobility shift assays revealed that PU.1 transactivated the OX40L promoter through direct binding to the most-proximal Ets motif. We found that this Ets motif is conserved between mouse and human, and that PU.1 bound to the human OX40L promoter in ChIP assay using human monocyte-derived DCs. ChIP assays based on ChIP-seq datasets revealed that PU.1 binds to several sites distant from the transcription start site on the OX40L gene in addition to the most-proximal site in mouse DCs. In the present study, the structure of the OX40L promoter regulated by PU.1 is determined. It is also suggested that PU.1 is involved in mouse OX40L expression via multiple binding sites on the gene.

Highlights

Despite the importance of OX40L in immune responses, the mechanism of the transcriptional regulation of OX40L gene remains to be elucidated
We found that PU.[1] binds to the Ets motif located in the 5′-flanking region proximal to the transcriptional start site and transactivates the OX40L gene both in mouse and human dendritic cells (DCs)
To evaluate the effect of PU.[1] suppression on OX40L expression, bone marrow-derived DCs (BMDCs) were transfected with PU.[1] small interfering RNA and stimulated with potent activators of DCs such as LPS, CpG, and poly I:C

Summary

Introduction

Despite the importance of OX40L in immune responses, the mechanism of the transcriptional regulation of OX40L gene remains to be elucidated. It was recently demonstrated that TSLP induces OX40L expression in DCs through the binding of NF-κB p50 and RelB to the OX40L promoter[23]. PU.[1] regulates gene expression by binding to canonical Ets motifs as a monomer and as a heterodimer with interferon regulatory factor 4 (IRF4) or IRF8, alternatively forming a complex with several transcription factors, including C/EBPαand β, and c-Jun[34]. We investigated whether PU.[1] regulates the expression of OX40L in DCs. We found that PU.[1] binds to the Ets motif located in the 5′-flanking region proximal to the transcriptional start site and transactivates the OX40L gene both in mouse and human DCs

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Conclusion