Upstream Distal Regulatory Elements Contact the Lmo2 Promoter in Mouse Erythroid Cells

Abstract

The Lim domain only 2 (Lmo2) gene encodes a transcriptional cofactor critical for the development of hematopoietic stem cells. Several distal regulatory elements have been identified upstream of the Lmo2 gene in the human and mouse genomes that are capable of enhancing reporter gene expression in erythroid cells and may be responsible for the high level transcription of Lmo2 in the erythroid lineage. In this study we investigate how these elements regulate transcription of Lmo2 and whether or not they function cooperatively in the endogenous context. Chromosome conformation capture (3C) experiments show that chromatin-chromatin interactions exist between upstream regulatory elements and the Lmo2 promoter in erythroid cells but that these interactions are absent from kidney where Lmo2 is transcribed at twelve fold lower levels. Specifically, long range chromatin-chromatin interactions occur between the Lmo2 proximal promoter and two broad regions, 3–31 and 66–105 kb upstream of Lmo2, which we term the proximal and distal control regions for Lmo2 (pCR and dCR respectively). Each of these regions is bound by several transcription factors suggesting that multiple regulatory elements cooperate in regulating high level transcription of Lmo2 in erythroid cells. Binding of CTCF and cohesin which support chromatin loops at other loci were also found within the dCR and at the Lmo2 proximal promoter. Intergenic transcription occurs throughout the dCR in erythroid cells but not in kidney suggesting a role for these intergenic transcripts in regulating Lmo2, similar to the broad domain of intergenic transcription observed at the human β-globin locus control region. Our data supports a model in which the dCR functions through a chromatin looping mechanism to contact and enhance Lmo2 transcription specifically in erythroid cells. Furthermore, these chromatin loops are supported by the cohesin complex recruited to both CTCF and transcription factor bound regions.

BRACHYURY directs histone acetylation to target loci during mesoderm development

“…One of these genes, Lmo2, is known to be an early upstream regulator of blood and endothelial development [40,41]. T binds to a region corresponding to the previously described À70 enhancer of Lmo2 (À70 kb from the TSS), which was shown by chromosome conformation capture assays to be in close proximity to the TSS of Lmo2 [42,43]. Both the À70 enhancer and the TSS of Lmo2 display a decrease in H3K27ac in T Y88A mutant mesodermal cells, in the ChIP-Seq data and confirmed by ChIP-qPCR ( Figs 5A and B, and EV5A).…”

Direct characterization of cis-regulatory elements and functional dissection of complex genetic associations using HCR–FlowFISH

“…We studied the moderately expressed gene LMO2 (79.8 TPM), a regulator of hematopoiesis for which multiple CREs have been previously nominated 27 , 28 . CASA identified a cluster of strong CREs +67–76 kb upstream of LMO2’s promoter ( Extended Data Fig.…”

“…4b ). These CREs reside within H3K27ac peaks that have been reported to interact directly with the LMO2 promoter and drive expression in vivo 27 , 28 . This stretch of CREs has been shown to recapitulate LMO2 expression in blood cells, consistent with our use of K562 cells 29 .…”

“Krüppeling Erythropoiesis”: KLF1 function in development and disease

“…These transcription factors include Gata1, Fog1, Nfe-2 and Tal1. 24,25,[36][37][38][39][40][41] Although there are shared transcription factors, it is likely that their target genes are specifically driven by lineage restricted transcription factors. Fli1 and Klf1 are unique and lineage restricted transcription factors that drive megakaryocytic and erythroid differentiation, respectively.…”