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. 2022 Aug 23;13:4941. doi: 10.1038/s41467-022-32566-9

Fig. 5. KLF4 modulates gene expression through changes in the enhancer landscape.

Fig. 5

a Scatterplot showing the relation between differential KLF4 binding at enhancer anchors identified by HiChIP, with target gene expression, in PAEC exposed to 15 dyn/cm2 of LSS vs ST conditions for 24 h. b Scatterplot showing the loss of enhancer anchors identified by HiChIP upon KLF siRNA (marked in red), and their impact on target gene expression. HiChIP enhancer anchors were derived from PAEC treated with KLF RNAi for 48 h prior to exposure to 15 dyn/cm2 of LSS vs ST conditions for an additional 24 h. c Scatterplot showing the relation between KLF4 binding at enhancer anchors identified by ABC, with target gene expression. n = 2 experimental replicates. d ABC, HiChIP and ChIP-Seq tracks illustrating a KLF4 bound enhancer loop targeting DUSP5, which is lost with KLF siRNA (left panel). DUSP5 mRNA was determined by RT-qPCR in PAEC exposed to 15 dyn/cm2 of LSS vs ST conditions for 24 h (top right panel). Silencing the enhancer by CRISPRi decreased DUSP5 mRNA levels (bottom right panel). e ABC, HiChIP, and ChIP-Seq tracks illustrating an enhancer loop targeting SMAD5, which was lost with KLF RNAi (left panel). SMAD5 mRNA was determined by RT-qPCR in PAEC exposed to 15 dyn/cm2 of LSS vs ST conditions for 24 h, and shown normalized to ST expression levels (top right panel). Silencing the enhancer by CRISPRi, decreased SMAD5 mRNA levels (bottom right panel). f ABC, HiChIP and ChIP-Seq tracks illustrating enhancer loops that target EDN1 under ST conditions, which are lost with LSS but are present with KLF siRNA (left panel). EDN1 mRNA was determined by RT-qPCR in PAEC exposed to 15 dyn/cm2 of LSS vs ST conditions for 24 h (top right panel). Silencing the enhancer increased EDN1 mRNA levels (bottom right panel). For a and b, N = 2 experimental replicates for KLF4 ChIP-Seq; n = 3 experimental replicates for HiChIP. For d–f, data are shown as the mean +/− s.e.m. For mRNA expression n = 5 experimental replicates; for CRISPRi experiments n = 3 replicates. P values were determined by Student’s two-tailed t-test. Source data are provided as a Source Data file.