We propose to study how the beta-globin locus control region (LCR) confers high-level transcriptional activity to the beta-globin genes over a long distance on a chromosome. Mechanisms of long-range transactivation are poorly understood.
In Specific Aim 1, we will define protein components of the LCR in living cells and will determine if they differ during erythroid differentiation. Despite extensive in vitro studies, little is known about the composition of native LCR complexes. We hypothesize that the components of the LCR vary during erythropoiesis to accommodate differential requirements for transactivation during development. A chromatin immunoprecipitation (ChIP) assay will be used to measure the binding of candidate proteins to the LCR.
In Specific Aim 2, we will define the histone acetylation and phosphorylation patterns of the beta-globin locus and will determine if the patterns change during erythroid differentiation. We hypothesize that the LCR recruits coactivators that establish a specific pattern of histone modifications throughout the beta-globin locus, which is necessary for long-range transactivation. We will define the pattern of histone modifications, will assess whether it changes during erythropoiesis, and will determine whether pharmacological inducers of fetal hemoglobin specifically modulate the pattern. We hypothesize that the histone modification pattern is established via primary and modulatory determinants.
In Specific Aim 3, we will identify primary determinants of the acetylation pattern. We have shown that the histone acetylase CBP/p300 is critical for LCR-mediated transactivation. We will test whether CBP/p300 is a primary determinant and will identify CBP/p300-interacting coactivators in erythroid cells. We will also assess whether specific histone deacetylases are primary determinants. The native structure of enhancer and LCR complexes and the histone modification pattern of a domain have not been defined in any system. Since long-range mechanisms control the transcription of multiple crucial genes that regulate cell proliferation and differentiation, our studies will yield principles of broad physiological and pathophysiological relevance. The long-term objective is to therapeutically modulate beta-globin gene expression in humans with hemoglobinopathies by perturbing specific steps of the mechanism by which the LCR regulate the beta-globin genes.
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| Liu, Jinhua; Li, Yapu; Tong, Jingyuan et al. (2018) Long non-coding RNA-dependent mechanism to regulate heme biosynthesis and erythrocyte development. Nat Commun 9:4386 |
| Katsumura, Koichi R; Mehta, Charu; Hewitt, Kyle J et al. (2018) Human leukemia mutations corrupt but do not abrogate GATA-2 function. Proc Natl Acad Sci U S A 115:E10109-E10118 |
| McIver, Skye C; Hewitt, Kyle J; Gao, Xin et al. (2018) Dissecting Regulatory Mechanisms Using Mouse Fetal Liver-Derived Erythroid Cells. Methods Mol Biol 1698:67-89 |
| Tanimura, Nobuyuki; Liao, Ruiqi; Wilson, Gary M et al. (2018) GATA/Heme Multi-omics Reveals a Trace Metal-Dependent Cellular Differentiation Mechanism. Dev Cell 46:581-594.e4 |
| Mehta, Charu; Johnson, Kirby D; Gao, Xin et al. (2017) Integrating Enhancer Mechanisms to Establish a Hierarchical Blood Development Program. Cell Rep 20:2966-2979 |
| Hewitt, Kyle J; Katsumura, Koichi R; Matson, Daniel R et al. (2017) GATA Factor-Regulated Samd14 Enhancer Confers Red Blood Cell Regeneration and Survival in Severe Anemia. Dev Cell 42:213-225.e4 |
| Katsumura, Koichi R; Bresnick, Emery H; GATA Factor Mechanisms Group (2017) The GATA factor revolution in hematology. Blood 129:2092-2102 |
| Liu, Peng; Sanalkumar, Rajendran; Bresnick, Emery H et al. (2016) Integrative analysis with ChIP-seq advances the limits of transcript quantification from RNA-seq. Genome Res 26:1124-33 |
| Gao, Xin; Wu, Tongyu; Johnson, Kirby D et al. (2016) GATA Factor-G-Protein-Coupled Receptor Circuit Suppresses Hematopoiesis. Stem Cell Reports 6:368-82 |
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