Our work focuses on the mechanisms by which transcription factors regulate the development of diverse hematopoietic lineages, and how these factors orchestrate terminal differentiation with cell cycle arrest. These functions require key nuclear factors to be able to both activate and repress transcription. GATA-1 activates as many genes as it represses during terminal erythroid differentiation in a manner largely dependent on the interaction with its co-factor FOG-1. We recently identified two novel mechanisms by which GATA-1 and FOG-1 regulate transcription. First, we purified a FOG-1-associated co-represser complex NuRD (nucleosome remodeling and deacetylase) that is required for GATA-1/FOG-1-mediated transcriptional repression. Second, using chromosome conformation capture (3C) we showed that GATA-1 and FOG-1 are required for the formation of long-range chromatin loops at the p-globin locus.
In Aim I we propose to study the function of the FOG-1-NuRD complex at the molecular and cellular level as well as in murine models. This includes the identification of the modules that mediate NuRD binding by FOG-1, determining their atomic structure, testing the function of NuRD subunits in erythroid cell lines, generating mice with mutations that disrupt the FOG-1-NuRD interaction, and the conditional knock out of the NuRD core subunit Mi-2p Since FOG-1 can function as co-activator and co-repressor for GATA-1, Aim II examines the mechanisms by which FOG-1 switches between these opposing functions. These studies include the analysis by CHIP and siRNA of NuRD components at activated and repressed genes.
This Aim will also investigate the role of posttranslational modifications of FOG-1 and NuRD with regard to the switch in activity. Based on encouraging preliminary results, in Aim III we will examine by 3C whether GATA-1 and FOG-1 actively form chromatin loops during the repression of the GATA-2 and c-kit genes. This requires a thorough prior characterization of the GATA-2 and c-kit loci with regard to GATA-1 binding and histone acetylation. The role of loop formation will be directly addressed in functional studies. We hope that together these studies will help to explain how key transcriptional regulators can switch between activating and repressive functions. This knowledge directly impacts on how nuclear proteins promote the formation of one lineage at the expense of another, and how they activate genes associated with the mature phenotype while simultaneously inhibiting a program that maintains the immature proliferative state. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK058044-06
Application #
7256430
Study Section
Erythrocyte and Leukocyte Biology Study Section (ELB)
Program Officer
Bishop, Terry Rogers
Project Start
2000-07-01
Project End
2010-06-30
Budget Start
2007-07-01
Budget End
2008-06-30
Support Year
6
Fiscal Year
2007
Total Cost
$328,441
Indirect Cost
Name
Children's Hospital of Philadelphia
Department
Type
DUNS #
073757627
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Dogan, Nergiz; Wu, Weisheng; Morrissey, Christapher S et al. (2015) Occupancy by key transcription factors is a more accurate predictor of enhancer activity than histone modifications or chromatin accessibility. Epigenetics Chromatin 8:16
Wu, Weisheng; Morrissey, Christapher S; Keller, Cheryl A et al. (2014) Dynamic shifts in occupancy by TAL1 are guided by GATA factors and drive large-scale reprogramming of gene expression during hematopoiesis. Genome Res 24:1945-62
Pimkin, Maxim; Kossenkov, Andrew V; Mishra, Tejaswini et al. (2014) Divergent functions of hematopoietic transcription factors in lineage priming and differentiation during erythro-megakaryopoiesis. Genome Res 24:1932-44
Deng, Wulan; Blobel, Gerd A (2014) Manipulating nuclear architecture. Curr Opin Genet Dev 25:1-7
Deng, Wulan; Rupon, Jeremy W; Krivega, Ivan et al. (2014) Reactivation of developmentally silenced globin genes by forced chromatin looping. Cell 158:849-860
Lai, Fan; Orom, Ulf A; Cesaroni, Matteo et al. (2013) Activating RNAs associate with Mediator to enhance chromatin architecture and transcription. Nature 494:497-501
Kadauke, Stephan; Udugama, Maheshi I; Pawlicki, Jan M et al. (2012) Tissue-specific mitotic bookmarking by hematopoietic transcription factor GATA1. Cell 150:725-37
Wang, Yuhuan; Meng, Ronghua; Hayes, Vincent et al. (2011) Pleiotropic platelet defects in mice with disrupted FOG1-NuRD interaction. Blood 118:6183-91
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
Lejon, Sara; Thong, Sock Yue; Murthy, Andal et al. (2011) Insights into association of the NuRD complex with FOG-1 from the crystal structure of an RbAp48·FOG-1 complex. J Biol Chem 286:1196-203

Showing the most recent 10 out of 33 publications