Cell fate choices by Tbx1 in forming the mammalian heart

Tbx1 在形成哺乳动物心脏过程中的细胞命运选择

基本信息

批准号：
10451598
负责人：
BERNICE E MORROW
金额：
$ 60.56万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-20 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10451598
关键词：
22q11 22q11.2 3-Dimensional APLN gene ATAC-seq Affect Alleles Anterior Aorta Binding Binding Proteins Binding Sites Biological Process Cardiac Cardiac development Cell Differentiation process Cell model Cells ChIP-seq Chromatin Chromosomes Data Data Set Defect DiGeorge Syndrome Ectopic Expression Embryo Embryonic Development Face Gene Expression Profile Gene Proteins General Population Genes Genetic Genetic Transcription Heart Heart Abnormalities Human Genetics Immunofluorescence Immunologic Individual Lung Mammals Maps Mesenchyme Mesoderm Mesoderm Cell Molecular Mus Muscle Muscle Fibers Mutant Strains Mice Mutation Myocardium Neck Neonatal Pathway interactions Patients Penetrance Persistent Truncus Arteriosus Pharyngeal Apparatus Phenocopy Population Population Sizes Process Regulatory Element Risk Factors Shprintzen syndrome Testing Time Tissues base cell behavior cell type conditional mutant congenital heart disorder embryo tissue experimental study functional genomics gastrulation gene network interest mouse model mutant precursor cell progenitor programs receptor single-cell RNA sequencing spatiotemporal stem cells transcription factor transcriptomics

项目摘要

TBX1 encodes a T-box transcription factor required for cardiac development. This gene maps to the chromosome 22q11.2 region that is deleted in patients with DiGeorge syndrome/velo-cardio- facial syndrome or 22q11.2 deletion syndrome (22q11.2DS). Approximately 60% of 22q11.2DS patients have congenital heart disease that mostly affects the cardiac outflow tract. A subset of individuals with a mutation of the TBX1 gene but not a deletion has been identified and they partially phenocopy patients with 22q11.2DS. Inactivation of one allele of Tbx1 in mice results in mild defects, but inactivation of both alleles results in neonatal lethality with a persistent truncus arteriosus, in which the aorta and pulmonary trunk fail to separate. This defect also occurs in 5- 10% of 22q11.2DS patients with congenital heart disease. To understand the function of Tbx1 in mammals, we performed single cell RNA-sequencing (scRNA-seq) of cardiopharyngeal mesoderm progenitor cells within the pharyngeal apparatus. We discovered a multilineage progenitor (MLP) population that expresses genes important for forming the cardiac outflow tract as well as branchiomeric muscles of the face and neck. The MLP population expands at the expense of more differentiated populations when Tbx1 is inactivated in the cardiopharyngeal mesoderm. Our main hypothesis is that Tbx1 is required in the MLP population for progression towards more differentiated states needed for cardiac development. We propose three specific aims to test this hypothesis. In the first aim, we will perform additional scRNA-seq experiments and analyze the complete dataset to understand how the progression of MLP cells are altered in Tbx1 conditional and global mutant mouse embryos. In Aim 2, we will determine where the MLP cells are localized in the embryo. Preliminary data suggests that these cells are localized to the nascent mesenchyme of the elongating pharyngeal apparatus. We will also inactivate Tbx1 specifically within the MLP cells to determine its particular functions. In Aim 3, we will turn to functional genomic studies and will identify open and accessible chromatin for which harbors TBX1 protein binding sites using ATAC-seq and ChIP-seq from embryo tissue. Preliminary data suggests that we are able to identify direct transcriptional target genes. By these three aims, we will understand the molecular functions of TBX1 in the progression of progenitor cells to more differentiated states to build the cardiac outflow tract.

TBX1 编码心脏发育所需的 T 盒转录因子。该基因映射到 DiGeorge 综合征/velo-cardio- 患者的染色体 22q11.2 区域被删除面部综合征或 22q11.2 缺失综合征 (22q11.2DS)。大约 22q11.2DS 的 60% 患者患有主要影响心脏流出道的先天性心脏病。的一个子集已鉴定出具有 TBX1 基因突变但未缺失的个体，并且他们具有 22q11.2DS 的部分表型患者。小鼠中 Tbx1 的一个等位基因失活会导致轻微缺陷，但两个等位基因失活会导致新生儿致死性持续性断干动脉瘤，其中主动脉和肺干无法分离。此缺陷也出现在 5- 10%的22q11.2DS患者患有先天性心脏病。了解Tbx1的功能哺乳动物，我们对心咽部进行了单细胞 RNA 测序 (scRNA-seq) 咽器内的中胚层祖细胞。我们发现了多谱系表达对形成心脏流出道重要的基因的祖细胞 (MLP) 群体以及面部和颈部的鳃肌。 MLP 群体在当 Tbx1 在心咽部失活时，更多分化人群的代价中胚层。我们的主要假设是 MLP 群体的进展需要 Tbx1 朝着心脏发育所需的更加分化的状态发展。我们提出三项具体建议旨在检验这一假设。在第一个目标中，我们将进行额外的 scRNA-seq 实验并分析完整的数据集，以了解 MLP 细胞的进展是如何改变的 Tbx1 条件突变和全局突变小鼠胚胎。在目标 2 中，我们将确定 MLP 的位置细胞定位于胚胎中。初步数据表明这些细胞位于伸长的咽器的新生间质。我们还将停用 Tbx1 特定于 MLP 细胞内以确定其特定功能。在目标 3 中，我们将转向功能基因组研究，并将确定开放且可访问的染色质使用 ATAC-seq 和 ChIP-seq 从胚胎组织中获得 TBX1 蛋白结合位点。初步数据表明我们能够识别直接转录靶基因。通过这三个目标，我们将了解 TBX1 在祖细胞进展到更多细胞过程中的分子功能不同的状态来构建心脏流出道。