Human diseases resulting from aberrant hemoglobin synthesis afflict large numbers of patients, and therapeutics for these disorders are far from optimal. Taken together with the merits of using the ?-globin locus as a model to understand cell type-specific transcriptional mechanisms, this problem has received considerable attention. Using innovative approaches, we identified factors and mechanisms controlling the adult ?-like globin genes. GATA-1 and its interactors function through dispersed sites at the ?-globin locus and loci encoding pivotal regulators of red cell biology, including heme biosynthetic enzymes. It is crucial to understand how factors select chromatin sites, recruit coregulators that modify/remodel chromatin, engage transcriptional machinery, and confer gene subnuclear localizations. Given powerful technologies, implicated factors, and unique expertise, we will address key issues to develop an integrative model to explain the transcriptional control of hemoglobin synthesis.
Specific Aim 1 - To test models to explain the relationship between mechanisms controlling globin genes and genes encoding heme biosynthetic enzymes. We hypothesize that GATA-1 functions with a unique cohort of trans-acting factors and coregulators to expel globin genes and the gene encoding a rate-limiting enzyme in heme biosynthesis, 4- aminolevulinic acid synthase (Alas2), from the nuclear periphery. This would establish a subnuclear positioning required for high-level transcription and coordinate globin chain production and heme synthesis. We will test models to explain how GATA-1-regulating factors initiate and/or maintain anchoring or repulsion of these loci at peripheral and internal sites.
Specific Aim 2 - To elucidate mechanisms underlying initiation and maintenance of GATA-1-dependent transcription. Nearly nothing is known about mechanisms that differentially initiate vs. maintain transcription, a problem particularly important for strategies to control transcription with a therapeutic outcome. We will use a powerful RNA interference asay to test the hypothesis that components regulating ?-globin and Alas2 transcription differ in their capacities to support initiation vs. maintenance.
Specific Aim 3 - To establish how GATA-1 mutations underlying human hematologic disorders dysregulate GATA factor-dependent genetic networks. Many questions remain unanswered regarding how disease mutations impair GATA-1 activity. We will determine how Arg216W and Arg216Gln mutations within the N-terminal zinc finger that underlie congenital erythropoietic porphyria, X- linked grey platelet syndrome, and thrombocytopenia with ?-thalassemia afect GATA-1 activity. These studies will yield broad insights into mechanisms controling hemoglobin synthesis, normal and malignant hematopoiesis, and additional developmental processes.

Public Health Relevance

The proposed studies shall provide key insights into mechanisms underlying disorders of hemoglobin synthesis, including thalassemias, anemia, hematopoiesis, hematologic malignancies, and diseases associated with aberrant heme biosynthesis, including porphyrias. The work shall also develop conceptual frameworks for the design of novel therapeutic strategies for these human disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37DK050107-16
Application #
8302334
Study Section
Molecular and Cellular Hematology (MCH)
Program Officer
Bishop, Terry Rogers
Project Start
1996-07-16
Project End
2016-07-31
Budget Start
2012-08-01
Budget End
2013-07-31
Support Year
16
Fiscal Year
2012
Total Cost
$322,988
Indirect Cost
$105,488
Name
University of Wisconsin Madison
Department
Pharmacology
Type
Schools of Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
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Katsumura, Koichi R; Yang, Chenxi; Boyer, Meghan E et al. (2014) Molecular basis of crosstalk between oncogenic Ras and the master regulator of hematopoiesis GATA-2. EMBO Rep 15:938-47
McIver, Skye C; Kang, Yoon-A; DeVilbiss, Andrew W et al. (2014) The exosome complex establishes a barricade to erythroid maturation. Blood 124:2285-97
DeVilbiss, Andrew W; Sanalkumar, Rajendran; Johnson, Kirby D et al. (2014) Hematopoietic transcriptional mechanisms: from locus-specific to genome-wide vantage points. Exp Hematol 42:618-29
Hewitt, Kyle J; Sanalkumar, Rajendran; Johnson, Kirby D et al. (2014) Epigenetic and genetic mechanisms in red cell biology. Curr Opin Hematol 21:155-64
Fujita, Rie; Takayama-Tsujimoto, Mariko; Satoh, Hironori et al. (2013) NF-E2 p45 is important for establishing normal function of platelets. Mol Cell Biol 33:2659-70
Pope, Nathaniel J; Bresnick, Emery H (2013) Establishment of a cell-type-specific genetic network by the mediator complex component Med1. Mol Cell Biol 33:1938-55
DeVilbiss, Andrew W; Boyer, Meghan E; Bresnick, Emery H (2013) Establishing a hematopoietic genetic network through locus-specific integration of chromatin regulators. Proc Natl Acad Sci U S A 110:E3398-407
Katsumura, Koichi R; DeVilbiss, Andrew W; Pope, Nathaniel J et al. (2013) Transcriptional mechanisms underlying hemoglobin synthesis. Cold Spring Harb Perspect Med 3:a015412
Chung, Dongjun; Kuan, Pei Fen; Li, Bo et al. (2011) Discovering transcription factor binding sites in highly repetitive regions of genomes with multi-read analysis of ChIP-Seq data. PLoS Comput Biol 7:e1002111

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