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Hewitt K.J.,University of Wisconsin - Madison | Hewitt K.J.,UW Madison Blood Research Program | Sanalkumar R.,University of Wisconsin - Madison | Sanalkumar R.,UW Madison Blood Research Program | And 6 more authors.
Current Opinion in Hematology | Year: 2014

PURPOSE OF REVIEW: Erythropoiesis, in which hematopoietic stem cells (HSCs) generate lineage-committed progenitors that mature into erythrocytes, is regulated by numerous chromatin modifying and remodeling proteins. We will focus on how epigenetic and genetic mechanisms mesh to establish the erythroid transcriptome and how studying erythropoiesis can yield genomic principles. RECENT FINDINGS: Trans-acting factor binding to small DNA motifs (cis-elements) underlies regulatory complex assembly at specific chromatin sites, and therefore unique transcriptomes. As cis-elements are often very small, thousands or millions of copies of a given element reside in a genome. Chromatin restricts factor access in a context-dependent manner, and cis-element-binding factors recruit chromatin regulators that mediate functional outputs. Technologies to map chromatin attributes of loci in vivo, to edit genomes and to sequence whole egnomes have been transformative in discovering critical cis-elements linked to human disease. SUMMARY: Cis-elements mediate chromatin-targeting specificity, and chromatin regulators dictate cis-element accessibility/function, illustrating an amalgamation of genetic and epigenetic mechanisms. Cis-elements often function ectopically when studied outside of their endogenous loci, and complex strategies to identify nonredundant cis-elements require further development. Facile genome-editing technologies provide a new approach to address this problem. Extending genetic analyses beyond exons and promoters will yield a rich pipeline of cis-element alterations with importance for red cell biology and disease. © 2014 Wolters Kluwer Health.

Hewitt K.J.,University of Wisconsin - Madison | Hewitt K.J.,UW Madison Blood Research Program | Kim D.H.,Seoul National University | Devadas P.,University of Wisconsin - Madison | And 15 more authors.
Molecular Cell | Year: 2015

Thousands of cis-elements in genomes are predicted to have vital functions. Although conservation, activity in surrogate assays, polymorphisms, and disease mutations provide functional clues, deletion from endogenous loci constitutes the gold-standard test. A GATA-2-binding, Gata2 intronic cis-element (+9.5) required for hematopoietic stem cell genesis in mice is mutated in a human immunodeficiency syndrome. Because+9.5 is the only cis-element known to mediate stem cell genesis, we devised a strategy to identify functionally comparable enhancers ("+9.5-like") genome-wide. Gene editing revealed+9.5-like activity to mediate GATA-2 occupancy, chromatin opening, and transcriptional activation. A+9.5-like element resided in Samd14, which encodes a protein of unknown function. Samd14 increased hematopoietic progenitor levels/activity and promoted signaling by a pathway vital for hematopoietic stem/progenitor cell regulation (stem cell factor/c-Kit), and c-Kit rescued Samd14 loss-of-function phenotypes. Thus, the hematopoietic stem/progenitor cell cistrome revealed a mediator of a signaling pathway that has broad importance for stem/progenitor cell biology. © 2015 Elsevier Inc.

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