Chromatin Structure, Epigenetic and Developmental Regulation of Mammalian Gene

National Institute Of Diabetes And Digestive And Kidney Diseases

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Abstract

The eukaryotic genome is organized at varying levels into chromosome territories, transcriptional compartments and topologically associating domains (TADs), which are architectural features largely shared between different cell types and across species. In contrast, within TADs, chromatin loops connect enhancers and their target genes to establish unique transcriptomes that distinguish cells and tissues from each other and underlie development and differentiation. We have been studying the widely expressed Lim domain binding 1 protein, LDB1, which we have shown plays a critical role in connecting enhancers and genes during hematopoiesis by forming a complex with erythroid cell-type specificity. GATA1, TAL1 and LMO2 are erythroid transcription factors that contribute to the chromatin looping and transcription activation properties of the LDB1 complex. Our recent work using erythroid progenitor cells and fetal liver cells shows that a repressor protein, ETO2, and an activator protein, hemogen, each participate in the LDB1 complex and recruit oppositely acting co-regulators to carry out epigenetic modulation of LDB1-binding enhancer. ETO2 recruits the repressive NuRD complex and hemogen recruits SWI/SNF remodeling complex. These two transcription factors create a toggle switch to regulate LDB1 complex function in gene expression during erythropoiesis. We have also carried out a genome wide CRISPR knock out screen (GecKO) to identify epigenetic novel factors and mechanisms required for erythropoiesis. The histone chaperone HIRA is a prominent candidate that we are actively investigating. LDB1 is a widely expressed and highly conserved protein from worms to humans. We recently found that LDB1 functions in embryonic stem cells are very limited but become important as cells are differentiated towards hematopoiesis. This has opened a window on the commissioning of the erythroid enhancer repertoire during differentiation of ESC to hematopoietic cells. LDB1 has long been known to have important roles in other developmental pathways such as neurogenesis, cardiogenesis and retinogenesis, although a mechanistic understanding of its role is lacking. By conditional knock out of LDB1 in liver we have uncovered a role for LDB1 in regulation of metabolic gene expression. Mechanistic studies in a human liver cell line further reveal collaboration of LDB1 with a cohort of transcription factors known to be important for liver gene regulation. We had previously observed that an enhancer RNA (eRNA), BGLT3, encoded within the human beta-globin locus, contributes to LDB1 long range chromatin looping in the locus, which activates globin gene transcription. We have now carried out a meta-analysis to identify globally eRNAs in erythroid cells that engage in long range interactions with gene promoters. We will use the candidates to address the question whether it is the eRNA per se, the locus or transcription of the locus that has a functional role in chromatin looping and gene expression. Overall, our studies are aimed at broadly and deeply understanding how LDB1 and co-regulators orchestrate changes in chromatin topology in diverse cell types in health and disease.