Long-range chromatin interactions control trophoblast-restricted HLA-G expression during pregnancy (IRM6P.657)

Abstract

Abstract Human pregnancy poses an immunological paradox: at the fetal-maternal interface, semi-allogeneic fetal extravillous trophoblasts (EVT) invade the uterine mucosa without being rejected by the maternal immune system. HLA-G is a nonclassical MHC class I molecule specifically expressed on the surface of EVT and is believed to be a key to fetus-induced immune tolerance. However, the EVT-specific expression of HLA-G is still poorly understood. A novel enhancer more than 10 kb upstream of HLA-G, Enhancer L, was discovered via dissection of the HLA-G locus using a Massively Parallel Reporter Assay (MPRA). DNase I Hypersensitive Site (DHS) mapping, Chromosome Conformation Capture (3C) and reporter gene assays established Enhancer L as a cell type-specific enhancer that loops into the HLA-G classical promoter. Strikingly, deletion of Enhancer L using a CRISPR/Cas9 dual guide approach results in complete ablation of HLA-G expression. Saturation mutagenesis of Enhancer L revealed the existence of motifs for transcription factors paramount in trophoblast development. Two of them, GATA2 and CEBPB, associate with Enhancer L and the HLA-G classical promoter, and are absolutely required for HLA-G expression. Interestingly, progesterone upregulates HLA-G, as well as GATA2, via a nonclassical membrane progesterone receptor complex in trophoblasts. These findings shed light on the mechanism by which HLA-G is specifically expressed in invasive trophoblasts during pregnancy.