A distant trophoblast-specific enhancer controls HLA-G expression at the maternal-fetal interface

作者：Ferreira Leonardo M R; Meissner Torsten B; Mikkelsen Tarjei S; Mallard William; O' Donnell Charles W; Tilburgs Tamara; Gomes Hannah A B; Camahort Raymond; Sherwood Richard I; Gifford David K; Rinn John L; Cowan Chad A; Strominger Jack L

来源：Proceedings of the National Academy of Sciences, 2016, 113(19): 5364-5369.

DOI：10.1073/pnas.1602886113

• human immune gene regulation
• pregnancy
• immune tolerance
• MPRA
• CRISPR/Cas9

HLA-G, a nonclassical HLA molecule uniquely expressed in the placenta, is a central component of fetus-induced immune tolerance during pregnancy. The tissue-specific expression of HLA-G, however, remains poorly understood. Here, systematic interrogation of the HLA-G locus using massively parallel reporter assay (MPRA) uncovered a previously unidentified cis-regulatory element 12 kb upstream of HLA-G with enhancer activity, Enhancer L. Strikingly, clustered regularly-interspaced short palindromic repeats (CRISPR)/Cas9-mediated deletion of this enhancer resulted in ablation of HLA-G expression in JEG3 cells and in primary human trophoblasts isolated fromplacenta. RNA-seq analysis demonstrated that Enhancer L specifically controls HLA-G expression. Moreover, DNase-seq and chromatin conformation capture (3C) defined Enhancer L as a cell type-specific enhancer that loops into the HLA-G promoter. Interestingly, MPRA-based saturation mutagenesis of Enhancer L identified motifs for transcription factors of the CEBP and GATA families essential for placentation. These factors associate with Enhancer L and regulate HLA-G expression. Our findings identify long-range chromatin looping mediated by core trophoblast transcription factors as the mechanism controlling tissue-specific HLA-G expression at the maternal-fetal interface. More broadly, these results establish the combination of MPRA and CRISPR/Cas9 deletion as a powerful strategy to investigate human immune gene regulation.

• 出版日期2016-5-10