The study of the epigenetic mechanisms that ensure the correct propagation of transcriptional states throughout the cell cycle is important for understanding lineage commitment and cellular reprogramming. The hematopoietic transcription factor GATA1, an acetylated protein, is essential for the differentiation and commitment of several hematopoietic lineages. We recently found that GATA1 interacts with the bromodomain protein Brd3 in an acetylation-dependent manner in vitro and in vivo. Anti-Brd3 ChIP-seq analysis showed that GATA1 is a key determinant of Brd3 recruitment in interphase cells in vivo. Notably, we found that Brd3, like some other Brd family proteins, remains bound to chromatin during mitosis when the vast majority of nuclear factors are removed from chromatin. Brd3 binding might thus provide an epigenetic memory function to "bookmark" genes in a manner that facilitates the appropriated re-assembly transcription factor complexes at the correct sites upon re-entry into the G1 phase of the cell cycle.
In Aim 1, we will study the function of Brd3 in hematopoietic development, and biochemically and structurally dissect the Brd3-GATA1 interaction. We will determine the in vivo acetylation sites of GATA1 and monitor acetylation dynamics during the cell cycle and at varying stages of cell growth and differentiation.
In Aim 2 we will examine the genome-wide occupancy pattern of Brd3 in mitosis and define the mechanisms of mitotic and interphase Brd3 recruitment. We will examine whether Brd3 serves a bookmarking function of hematopoietic genes and study how Brd3 transmits transcriptional information through mitosis. This will involve the testing of Brd3-occupied elements in gain-of- function assays at the single allele level in living cells and the targeted destruction of Brd3 specifically during mitosis.
In Aim 3 the in vivo function of Brd3 will be assessed by conditional deletion in mice. Together, the proposed studies are designed to elucidate the transcriptional and epigenetic machinery that underlies the establishment and stable propagation of gene expression patterns in the hematopoietic system.

Public Health Relevance

The proposed studies are aimed to better understand the mechanisms underlying the formation of the blood lineages and their associated disorders. Our work focuses on how nuclear factors and their chemical modifications control the genes that govern the specification and differentiation of blood cells. Progress in this area promises novel approaches to treat blood-related disorders including leukemias and anemias.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK054937-15
Application #
8467704
Study Section
Molecular and Cellular Hematology (MCH)
Program Officer
Bishop, Terry Rogers
Project Start
1999-03-01
Project End
2016-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
15
Fiscal Year
2013
Total Cost
$351,562
Indirect Cost
$141,674
Name
Children's Hospital of Philadelphia
Department
Type
DUNS #
073757627
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Hardison, Ross C; Blobel, Gerd A (2013) Genetics. GWAS to therapy by genome edits? Science 342:206-7
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Lamonica, Janine M; Deng, Wulan; Kadauke, Stephan et al. (2011) Bromodomain protein Brd3 associates with acetylated GATA1 to promote its chromatin occupancy at erythroid target genes. Proc Natl Acad Sci U S A 108:E159-68
Gamsjaeger, Roland; Webb, Sarah R; Lamonica, Janine M et al. (2011) Structural basis and specificity of acetylated transcription factor GATA1 recognition by BET family bromodomain protein Brd3. Mol Cell Biol 31:2632-40
Wu, Weisheng; Cheng, Yong; Keller, Cheryl A et al. (2011) Dynamics of the epigenetic landscape during erythroid differentiation after GATA1 restoration. Genome Res 21:1659-71
Miccio, Annarita; Blobel, Gerd A (2010) Role of the GATA-1/FOG-1/NuRD pathway in the expression of human beta-like globin genes. Mol Cell Biol 30:3460-70
Bersenev, Alexey; Wu, Chao; Balcerek, Joanna et al. (2010) Lnk constrains myeloproliferative diseases in mice. J Clin Invest 120:2058-69
Deng, Wulan; Blobel, Gerd A (2010) Do chromatin loops provide epigenetic gene expression states? Curr Opin Genet Dev 20:548-54

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