Transcriptional Control of Maternal-Fetal Immune Tolerance

Human leukocyte antigens (HLA) are important determinants of self-nonself immune recognition. HLA-G, uniquely expressed in the placenta, is believed to be key to fetus-induced immune tolerance during pregnancy. The tissue-specific expression of HLA-G, however, remains poorly understood. Using a Massively Parallel Reporter Assay (MPRA), we discovered a 121 bp sequence 12 kb upstream of HLA-G with enhancer activity, Enhancer L. Strikingly, deletion of Enhancer L using a CRISPR/Cas9 dual guide approach resulted in complete ablation of HLA-G expression in a trophoblast cell line. This finding was confirmed in primary extravillous trophoblasts isolated from human placenta. RNA-seq analysis demonstrated that Enhancer L regulates HLA-G expression specifically. Moreover, DNase-seq and Chromatin Conformation Capture (3C) defined Enhancer L as a cell type-specific enhancer that loops into the HLA-G promoter. GATA2, GATA3, and CEBPB, factors essential for placentation, associate with Enhancer L and regulate HLA-G expression levels. These results establish long-range chromatin looping as a novel mechanism controlling trophoblast-specific HLA-G expression at the maternal-fetal interface.Biology, Molecular and Cellula