Abstract
Identification of cell type-specific enhancers is important for understanding the regulation of programs controlling cellular development and differentiation. Enhancers are typically marked by the co-transcriptional activator protein p300 or by groups of cell-expressed transcription factors. We hypothesized that a unique set of enhancers regulates gene expression in human erythroid cells, a highly specialized cell type evolved to provide adequate amounts of oxygen throughout the body. Using chromatin immunoprecipitation followed by massively parallel sequencing, genome-wide maps of candidate enhancers were constructed for p300 and four transcription factors, GATA1, NF-E2, KLF1, and SCL, using primary human erythroid cells. These data were combined with gene expression analyses, and candidate enhancers were identified. Consistent with their predicted function as candidate enhancers, there was statistically significant enrichment of p300 and combinations of co-localizing erythroid transcription factors within 1-50 kb of the transcriptional start site (TSS) of genes highly expressed in erythroid cells. Candidate enhancers were also enriched near genes with known erythroid cell function or phenotype. Candidate enhancers exhibited moderate conservation with mouse and minimal conservation with nonplacental vertebrates. Candidate enhancers were mapped to a set of erythroid-associated, biologically relevant, SNPs from the genome-wide association studies (GWAS) catalogue of NHGRI, National Institutes of Health. Fourteen candidate enhancers, representing 10 genetic loci, mapped to sites associated with biologically relevant erythroid traits. Fragments from these loci directed statistically significant expression in reporter gene assays. Identification of enhancers in human erythroid cells will allow a better understanding of erythroid cell development, differentiation, structure, and function and provide insights into inherited and acquired hematologic disease.
Highlights
Programs of cellular development and differentiation are controlled by enhancers
An alternative way to predict the presence of cis-regulatory modules (CRMs) is the regulatory potential (RP) score, which evaluates whether regions of DNA sequence have patterns more similar to those of Evolutionary conservation at sites of p300 and erythroid transcription factor occupancy in distal, intergenic, and intron regions in erythroid cell chromatin Conservation of human candidate enhancer regions was analyzed using the UCSC LiftOver tool at stringency levels of 75 or 50% of bases in the region that must remap
Mammalian erythroid cells are an excellent example of the complexity in temporal, developmental, and differentiation stage-specific changes exhibited by a single cell type
Summary
Programs of cellular development and differentiation are controlled by enhancers. Results: Human erythroid cell type-specific enhancers are marked by p300 and groups of transcription factors. Using chromatin immunoprecipitation followed by massively parallel sequencing, genome-wide maps of candidate enhancers were constructed for p300 and four transcription factors, GATA1, NF-E2, KLF1, and SCL, using primary human erythroid cells. Identification and characterization of enhancers that control programs of gene expression in highly specialized human erythroid cells will allow a better understanding of erythroid cell development, differentiation, structure, and function as well as provide insights into inherited and acquired hematologic disease. Genome-wide maps of p300 and four erythroid transcription factors, GATA1, NF-E2, KLF1, and SCL, in human primary erythroid cell chromatin were constructed and analyzed with parallel gene expression analyses Consistent with their predicted function, these regulatory elements were enriched near genes highly expressed in erythroid cells or involved in erythroid cell structure and function. Fragments from 9 of the 10 biologically relevant regions directed statistically significant expression in reporter gene assays
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