In metazoans, gene expression is regulated in a tissue/cell-type specific manner predominantly via stretches of noncoding sequence referred to as cis regulatory modules (CRMs) that regulate the expression of (typically) the adjacent gene(s). CRMs usually contain 1 or more DNA binding sites for 1 or more sequence-specific, regulatory transcription factors (TFs) that function to activate or repress the target gene(s);CRMs that activate gene expression are frequently referred to as ?transcriptional enhancers?, and have been the focus of many computational and experimental studies. Identification of tissue/cell-type-specific enhancers in metazoans remains a significant challenge. Moreover, despite recent technological advances, a major, rate-limiting bottleneck that is impeding rapid progress in the field is the still quite low-throughput experimental testing of candidate enhancers. The overarching goals of this project are to develop and apply novel ?wet-lab? technology for high-throughput experimental identification of tissue/cell-type-specific transcriptional enhancers. In this project we will focus on the developing embryonic mesoderm in Drosophila as a model system. We will identify cis regulatory modules and analyze their constituent cis regulatory codes that operate in somatic mesoderm (SM) founder cells (FCs) and fusion competent myoblasts (FCMs). Specifically, we will: develop and apply novel ?wet-lab? technology for high-throughput experimental identification of tissue/cell-type-specific transcriptional enhancers;determine the DNA binding specificities of ~75 known and predicted TFs expressed in the Drosophila embryonic mesoderm;predict CRMs and infer cis regulatory codes considering highly combinatorial input from large, high-resolution TF-DNA binding specificity dictionaries;and experimentally validate newly discovered enhancers. Importantly, we anticipate that the technologies, approaches, tools, and data resulting from this project will be generally applicable to other systems and organisms.

Public Health Relevance

In metazoans, gene expression is regulated in a tissue/cell-type specific manner predominantly via stretches of noncoding sequence referred to as cis regulatory modules. The overarching goals of this renewal project are to develop and apply novel 'wet-lab'technologies for high-throughput experimental identification of tissue/cell-type- specific transcriptional enhancers, and to use them to decipher cis regulatory codes that control gene expression in a tissue- and cell-type specific manner. In this project we will focus on the developing embryonic mesoderm in Drosophila as a model system. There is remarkable conservation of all the major regulatory components, including both signals and TFs, governing heart and muscle development in vertebrates and Drosophila. A deeper understanding of these pathways and their integration is essential for developing rational approaches to congenital heart disease and muscular dystrophies in children and to cardiac regeneration for acquired heart disorders in adults. We anticipate that the technologies, approaches, tools, and data resulting from this project will be generally applicable to other systems and organisms.

Agency
National Institute of Health (NIH)
Institute
National Human Genome Research Institute (NHGRI)
Type
Research Project (R01)
Project #
5R01HG005287-03
Application #
8312733
Study Section
Special Emphasis Panel (ZRG1-GGG-M (91))
Program Officer
Pazin, Michael J
Project Start
2010-09-27
Project End
2013-10-31
Budget Start
2012-08-01
Budget End
2013-10-31
Support Year
3
Fiscal Year
2012
Total Cost
$569,758
Indirect Cost
$250,566
Name
Brigham and Women's Hospital
Department
Type
DUNS #
030811269
City
Boston
State
MA
Country
United States
Zip Code
02115
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