Mapping Transcriptional Networks in Cardiac Development

绘制心脏发育中的转录网络

• 批准号: 7936097
• 负责人: JONATHAN G SEIDMAN
• 金额: $ 194.76万
• 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/7936097
• 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; biochip; congenital heart disorder; genome-wide; human data; insight; malformation; new technology; novel; novel therapeutic intervention; promoter; research study; transcription factor

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）： 心脏发展是由多种监管过程产生的。 HCCB研究CTR的重点是定义这些过程之一 - 转录调节网络。 RNA转录的调节被认为在发育过程中起关键作用，对转录调节剂的整个曲目的定义及其在心脏胚胎发生过程中的相互作用仍然很大未知。尽管已经鉴定出大量的转录因子与心脏发展有关，但激活这些转录因子的过程以及这些转录因子的下游目标尚不清楚。我们建议采用新型的基因组和遗传方法来定义转录因子网络，这些转录因子网络调节心脏发育中的多个步骤并覆盖该网络在这些发育中的组织中发现的RNA表达谱上。最后，这应该为理解心脏发展提供RNA蓝图。心脏发展所需的网络的定义将增强对最近发现的后调节机制（例如microRNA）的作用的理解，这些机制可能会改变发展过程。我们将利用新型技术来定义RNA表达网络，包括基因表达的时间和空间模式，这些基因表达形成了原发性和次要心脏场与成人腔室形成。将采用三种不同的新技术来定义每个组织中的RNA谱在临界的临界阶段（DSAGE和RNASEQ），并确定可能的特定转录因子（BioChip）的可能下游靶标。 DSAGE和RNASEQ采用高吞吐量DNA测序技术来定义RNA表达的结构和水平，而生物芯片则定义了受特定转录因子绑定的基因组中的启动子位点。这些信息将共同定义转录因子水平及其下游靶基因。被确定为发育步骤中心的分子将通过扰动实验（包括分子和表型分析）来验证。与人类研究的数据一起，我们将构建心脏发育的蓝图，并询问这种蓝图的扰动如何导致先天性心脏病。 1）在使用DSAGE和RNASEQ的一级心脏场，次生心脏场和心脏室中定义转录组。 2）确定使用Dchip使用DCHIP的GATA4，F0G2，TBX5，NKX2.5的结合靶标和其他对于心脏发育至关重要的转录因子。 3）构建一种发育蓝图，用于从.model有机体和人类数据集中的心脏规范和形态发生的发育阶段。 4）通过扰动RNA水平并定义分子和表型后果来验证定义蓝图中淋巴结的贡献作用。相关性（请参阅说明）：拟议研究的目的是定义导致成熟心脏形成的转录网络。定义该网络将为各种先天性心脏病（CHD）的发病机理提供新的见解。最终，此类研究可能会允许新的治疗方法来验证CHD。