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. 2019 Jun 28;47(16):8424–8438. doi: 10.1093/nar/gkz560

Figure 6.

Figure 6.

A distant-acting enhancer contacts the promoter region of ENPP2 and controls its expression. (A) Circos plot showing interactions between ENPP2 TSS and a putative enhancer located at 82 kb, dataset from CD34+ cells. (B) 3C experiments replicating this interaction in HEK 293T cells. LPS treatment did not modulate this interaction. (C) H3K4me1 enrichment at the enhancer region, as measured by ChIP-qPCR, using six sets of primers covering the putative enhancer region. (D) Luciferase reporter assay demonstrating that the region has enhancer activity, independently of its orientation. (E) Scheme depicting the deletion of the enhancer region by using the CRISPR-Cas9 technology. Cas9 enzyme was transfected simultaneously with two gRNA targeting 5′ end and 3′ end of the enhancer region. (F) Endogenous expression of ENPP2 after CRISPR-Cas9-mediated deletion of the enhancer, as measured by q-RT-PCR. LPS 100 ng/ml.