Mapping Transcriptional Networks in Cardiac Development

绘制心脏发育中的转录网络

• 批准号: 8663738
• 负责人: JONATHAN G SEIDMAN
• 金额: $ 1.58万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8663738
• 关键词: Adult; Animal Model; Binding; Biotin; Cardiac; Cataloging; Catalogs; Cells; Chickens; DNA Sequence; Data; Data Set; Development; Developmental Process; Embryonic Development; Gene Expression; Gene Expression Profile; Gene Targeting; Genes; Genetic; Genetic Transcription; Genome; Genomics; Goals; Heart; Heart Diseases; Human; Instruction; Lead; Location; Maps; Methods; MicroRNAs; Molecular; Molecular Profiling; Morphogenesis; Mus; Pathogenesis; Pathway interactions; Pattern; Phosphorylation; Play; Process; Proteins; RNA; RNA Sequences; Regulation; Regulatory Element; Research; Role; Sequence Analysis; Site; Staging; Structure; Technology; Tissues; Transcript; Tube; activating transcription factor; congenital heart disorder; deep sequencing; genome-wide; human data; insight; malformation; new technology; novel; novel therapeutic intervention; promoter; research study; transcription factor; transcriptome sequencing

DESCRIPTION (provided by applicant): Cardiac development results from a variety of regulatory processes. The HCCB Research Ctr is focused on defining one of these processes-the transcriptional regulatory network. Regulation of RNA transcription is recognized to play critical roles in developmental processes, definition of the entire repertoire of transcriptional regulators and their interactions during cardiac embryogenesis remains largely unknown. While large numbers of transcription factors have been identified that are involved in cardiac development the processes that activate these transcription factors and the downstream targets of these transcription factors are not known. We propose to employ novel genomic and genetic methods to define the network of transcription factors that regulate multiple steps in cardiac development and overlay that network on the RNA expression profile found in these developing tissues. In the end this should provide an RNA blueprint for understanding cardiac development. Definition of the network required for cardiac development would enhance understandings of the roles of recently discovered post-transcriptional regulatory mechanisms (e.g. microRNAs) that may modify developmental processes. We will take advantage of novel technologies to define the RNA expression networks, including both temporal and spatial patterns of gene expression that formation of primary and secondary heart fields to adult chamber formation. Three different novel technologies will be employed to define RNA profiles in each tissue at critical stages of development (DSAGE and RNAseq) and to identify likely downstream targets of particular transcription factors (bioCHIP). DSAGE and RNAseq employ high throughput DNA sequencing technologies to define the structure and level of RNA expression, while bioCHIP defines the promoter sites within the genome that are bound by particular transcription factors. Together this information will define transcription factor levels and their downstream target genes. Molecules that are identified as central for developmental steps will be validated by perturbation experiments, including molecular and phenotypic analyses. Together with data from human studies, we will construct a cardiac developmental blueprint and interrogate how perturbations of this blueprint result in congenital heart disease. 1) Define the transcriptome in the primary heart field, the secondary heart field and in developing cardiac chambers using DSAGE and RNAseq. 2) Identify binding targets of GATA4, F0G2, TBX5, NKX2.5 and other transcription factors critical for cardiac development using DCHIP. 3) Construct a developmental blueprint for developmental stages of cardiac specification and morphogenesis from .model organism and human datasets. 4) Validate crifical role of defines nodes in the blueprint by perturbing RNA levels and defining molecular and phenotypic consequences. RELEVANCE (See instructions): The goal of the proposed research is to define transcriptional networks that lead to formation of the mature heart. Defining this network will provide novel insights into the pathogenesis of a variety of congenital heart diseases (CHD). Eventually such studies may allow new therapeutic approaches to CHD.

描述（由申请人提供）： 心脏发育是由多种调节过程引起的。HCCB研究中心的重点是定义这些过程之一-转录调控网络。RNA转录调控在发育过程中起着重要的作用，但在心脏胚胎发生过程中，转录调控因子及其相互作用的定义仍不清楚。虽然已经鉴定了大量参与心脏发育的转录因子，但是激活这些转录因子的过程和这些转录因子的下游靶点尚不清楚。我们建议采用新型基因组和遗传方法来定义调节心脏发育多个步骤的转录因子网络，并将该网络覆盖在这些发育组织中发现的RNA表达谱上。最终，这将为理解心脏发育提供一个RNA蓝图。心脏发育所需网络的定义将增强对最近发现的可能改变发育过程的转录后调节机制（例如微小RNA）的作用的理解。我们将利用新的技术来定义RNA表达网络，包括基因表达的时间和空间模式，即从初级和次级心脏领域到成人心室形成。将采用三种不同的新技术来定义发育关键阶段每个组织中的RNA谱（DSAGE和RNAseq），并确定特定转录因子的可能下游靶点（bioCHIP）。DSAGE和RNAseq采用高通量DNA测序技术来定义RNA表达的结构和水平，而bioCHIP则定义了基因组内与特定转录因子结合的启动子位点。这些信息将共同定义转录因子水平及其下游靶基因。将通过扰动实验（包括分子和表型分析）验证鉴定为发育步骤中心的分子。结合人类研究的数据，我们将构建一个心脏发育蓝图，并询问这个蓝图的扰动如何导致先天性心脏病。1)使用DSAGE和RNAseq定义主要心脏区域、次要心脏区域和发育中的心腔中的转录组。2)使用DCHIP鉴定GATA 4、F0 G2、TBX 5、NKX2.5和其他对心脏发育至关重要的转录因子的结合靶点。3)从模式生物和人类数据集构建心脏规格和形态发生发育阶段的发育蓝图。4)通过扰乱RNA水平并定义分子和表型结果，定义蓝图中的节点。相关性（见说明）：拟议研究的目标是定义导致成熟心脏形成的转录网络。定义这个网络将提供新的见解，各种先天性心脏病（CHD）的发病机制。最终，这些研究可能会为CHD提供新的治疗方法。