Krϋppel-like factor-1 (KLF1) is an essential transcription factor and regulator of gene expression during erythropoiesis. A number of studies have shown nearly 1000 genes are poorly expressed when KLF1 is absent. This global loss of expression is responsible for failure of effective red blood cell production in KLF1 knockout mice, and partly responsible for the anemia in humans with dominant mutations in KLF1. To determine whether these KLF1-dependent genes are direct or indirect targets of KLF1, we have previously performed global ChIP-seq experiments identifying 945 regions of KLF1 occupancy in the mouse genome. About 15% of these regions fall within the promoters of KLF1 target genes but surprisingly, most are thousands of kilobases distant from any known gene. Yet, many of these distant sites still exhibit co-occupancy with other transcriptional regulators involved in erythropoiesis, including GATA1 (GATA binding protein 1) and within regions displaying enhancer signatures (for example H3K4me1). The nature and function of these sites is interesting as they may shed new light onto how KLF1 and other associated transcription factors really work in cells.

To explore the function of these KLF1-bound loci, we have performed chromosome conformation capture coupled with next-generation sequencing (Capture-C) using a tamoxifen responsive, KLF1 inducible cell line to investigate the role of KLF1 in chromosomal looping. In combination with ATAC-seq and KLF1 ChIP-seq in these cells, we have analyzed the primary transcriptional output of KLF1 target genes, including β-globin, a-globin and Dematin to assess the transcriptional response to KLF1-depedent enhancer-promoter interactions. We have identified multiple loci genome-wide where occupancy of KLF1 is critical for promoter-enhancer interactions leading to transcriptional regulation of distal target genes.