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Insight into GATA1 transcriptional activity through interrogation of cis elements disrupted in human erythroid disorders

Item Type:Article
Title:Insight into GATA1 transcriptional activity through interrogation of cis elements disrupted in human erythroid disorders
Creators Name:Wakabayashi, A. and Ulirsch, J.C. and Ludwig, L.S. and Fiorini, C. and Yasuda, M. and Choudhuri, A. and McDonel, P. and Zon, L.I. and Sankaran, V.G.
Abstract:Whole-exome sequencing has been incredibly successful in identifying causal genetic variants and has revealed a number of novel genes associated with blood and other diseases. One limitation of this approach is that it overlooks mutations in noncoding regulatory elements. Furthermore, the mechanisms by which mutations in transcriptionalcis-regulatory elements result in disease remain poorly understood. Here we used CRISPR/Cas9 genome editing to interrogate three such elements harboring mutations in human erythroid disorders, which in all cases are predicted to disrupt a canonical binding motif for the hematopoietic transcription factor GATA1. Deletions of as few as two to four nucleotides resulted in a substantial decrease (>80%) in target gene expression. Isolated deletions of the canonical GATA1 binding motif completely abrogated binding of the cofactor TAL1, which binds to a separate motif. Having verified the functionality of these three GATA1 motifs, we demonstrate strong evolutionary conservation of GATA1 motifs in regulatory elements proximal to other genes implicated in erythroid disorders, and show that targeted disruption of such elements results in altered gene expression. By modeling transcription factor binding patterns, we show that multiple transcription factors are associated with erythroid gene expression, and have created predictive maps modeling putative disruptions of their binding sites at key regulatory elements. Our study provides insight into GATA1 transcriptional activity and may prove a useful resource for investigating the pathogenicity of noncoding variants in human erythroid disorders. SIGNIFICANCE: Methods for identifying causal variants underlying human diseases have been greatly enhanced by whole-exome sequencing; however, this approach overlooks mutations that occur within noncoding regulatory regions. Moreover, the mechanisms for how such mutations result in disease are poorly understood. In this study, we interrogated binding sites of the blood cell transcription factor GATA1 in regulatory elements that are mutated in cases of human red blood cell disorders by creating small-targeted deletions in model cell lines. These deletions cause a major reduction in target gene expression. We used this initial insight to show that such elements are highly conserved, and that through predictive modeling, we can gain global insight into key determinants of GATA1 transcriptional activity.
Keywords:GATA1, Cis-Regulatory Elements, Noncoding Mutations, Mendelian Erythroid Disorders
Source:Proceedings of the National Academy of Sciences of the United States of America
Publisher:National Academy of Sciences
Page Range:4434-4439
Date:19 April 2016
Official Publication:https://doi.org/10.1073/pnas.1521754113
External Fulltext:View full text on PubMed Central
PubMed:View item in PubMed

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