This project is to use high-throughput genomic sequencing analysis of histone marks and transcription factor binding sites to track the succession of regulatory changes that underlie multipotent progenitor differentiation into T lymphocytes. The project will characterize the transitions from one stage to the next in terms of genome-wide changes in histone modifications (""""""""epigenetic marks""""""""), changes in RNA polymerase II loading and activity, and changes in comprehensively measured transcript accumulation. The feasibility of this project is based not only on access to extensive Solexa/Illumina sequencing and quantitative informatics analysis, but also on the advent of efficient methods for expanding and isolating primary mouse hematopoietic cells at a series of discrete phases across the transition from stem cell to committed T cell precursor. This system provides unusually advantageous access to cell states in a real, precisely coordinated developmental continuum. As defined by previous work of the applicants, the T-cell developmental sequence includes opportunities to observe both the mechanisms through which T lineage-specific genes are first activated and the mechanisms through which stem cell pluripotency genes and self-renewal genes are programmed for long- term silencing. The project thus offers a novel opportunity for the epigenomics field to test the significance of patterns of epigenetic marking at sites throughout the genome, in light of known trajectories of changing expression of these loci over time.

Public Health Relevance

The genome that is the blueprint for life includes much more than sequences that code directly for proteins. It also includes sequences that are control sites where levels of RNA and protein expression can be regulated. As stem cells develop into mature cell types, for example the blood cells that are produced every day throughout a human life, thousands of genes must be re-tuned in their expression levels to create the new cell identity. This regulation can be extremely sensitive, as small variations can lead to disease. Recent advances help to map where regulatory sequence sites are likely to lie in mammalian genomes, but knowledge of the rules that govern when they are acting have so far lagged behind. What is needed is to bring these new technologies to bear on a well characterized set of mammalian cells caught at various stages in the process of development from a stem cell to a differentiated fate, where the changes across the genome can be followed across time. In this project, we therefore use advanced technology for tracking the action at regulatory sequences as differentiation progresses to understand in detail the genome-wide events that cause a stem cell to develop into a T lymphocyte of the immune system. This system is profoundly important to human health. In this project, we will determine how different types of regulatory events occurring across the genome can explain which type of cell the precursors become, as well as how they avoid being sidetracked into leukemia. .

National Institute of Health (NIH)
National Cancer Institute (NCI)
High Impact Research and Research Infrastructure Programs (RC2)
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Special Emphasis Panel (ZCA1-RPRB-O (O9))
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Mccarthy, Susan A
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California Institute of Technology
Schools of Arts and Sciences
United States
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Rothenberg, Ellen V (2017) Fitting structure to function in gene regulatory networks. Hist Philos Life Sci 39:37
Longabaugh, William J R; Zeng, Weihua; Zhang, Jingli A et al. (2017) Bcl11b and combinatorial resolution of cell fate in the T-cell gene regulatory network. Proc Natl Acad Sci U S A 114:5800-5807
Rothenberg, Ellen V; Kueh, Hao Yuan; Yui, Mary A et al. (2016) Hematopoiesis and T-cell specification as a model developmental system. Immunol Rev 271:72-97
Kueh, Hao Yuan; Yui, Mary A; Ng, Kenneth K H et al. (2016) Asynchronous combinatorial action of four regulatory factors activates Bcl11b for T cell commitment. Nat Immunol 17:956-65
Rothenberg, Ellen V (2015) Immune Cell Identity: Perspective from a Palimpsest. Perspect Biol Med 58:205-28
Scripture-Adams, Deirdre D; Damle, Sagar S; Li, Long et al. (2014) GATA-3 dose-dependent checkpoints in early T cell commitment. J Immunol 193:3470-91
Rothenberg, Ellen V (2014) The chromatin landscape and transcription factors in T cell programming. Trends Immunol 35:195-204
Rothenberg, Ellen V (2014) Transcriptional control of early T and B cell developmental choices. Annu Rev Immunol 32:283-321
Li, Long; Zhang, Jingli A; Dose, Marei et al. (2013) A far downstream enhancer for murine Bcl11b controls its T-cell specific expression. Blood 122:902-11
Rothenberg, Ellen V; Champhekar, Ameya; Damle, Sagar et al. (2013) Transcriptional establishment of cell-type identity: dynamics and causal mechanisms of T-cell lineage commitment. Cold Spring Harb Symp Quant Biol 78:31-41

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