Fig. S3.

Enhancer L loops into the classical promoter of HLA-G. (A) Strategy for chromatin conformation capture (3C) analysis of the HLA-G locus. DpnII restriction sites flank the two regions of interest: Enhancer L, labeled in red, and the HLA-G classical promoter, given in blue. Primers in black amplify a product that serves as “loading control,” and primers in red and in blue were used to detect loop formation. (B) 3C indicates that Enhancer L physically interacts with the classical promoter of HLA-G in JEG3, but not in HLA-G negative HEK293T cells. C, Loading control PCR product (322 bp); L, Enhancer L–classical promoter looping interaction PCR product (640 bp), marked with a star. (C) Schematic representing the main steps in 3C: chromatin cross-linking, restriction digest, ligation at low DNA concentrations, and PCR-based detection of looping interactions. (D) Sequence confirmation of the physical interaction between Enhancer L and the classical promoter of HLA-G detected by 3C assays (PCR amplicon marked with a star in Fig. S3B). Primer pointing away from the classical promoter is depicted in blue, primer pointing away from Enhancer L is in red, and the DpnII restriction site where ligation of the contact regions occurred is enclosed by a gray-shaded box.