The Role of YAP1:TEAD Enhancers in the Retinoic Acid-Regulatory Network of Cardiac Progenitors

YAP1:TEAD 增强剂在心脏祖细胞视黄酸调节网络中的作用

• 批准号: 10683794
• 负责人: Concepcion Estaras
• 金额: $ 46.87万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-12 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10683794
• 关键词: 22q11.2; ATAC-seq; Address; Adopted; Affect; Binding; Binding Proteins; Biological Assay; Biopsy; Cardiac; Cardiac Myocytes; Cardiac development; Cause of Death; Cell Differentiation process; Cell Fate Control; Cells; ChIP-seq; Chromatin; Collaborations; Complex; Congenital Heart Defects; Cues; Data; Defect; Development; DiGeorge Syndrome; Echocardiography; Enhancers; Exposure to; Frequencies; Genes; Genetic; Genetic Transcription; Genomic approach; Genotype; Goals; Heart; Heart Atrium; Histopathology; Homeostasis; Human; In Vitro; Incidence; Infant; Knowledge; Lead; Molecular; Mus; Mutation; Nuclear Receptors; Nutritional; Pathway Analysis; Pathway interactions; Patients; Phenotype; Phosphotransferases; Population; Property; Recovery; Resolution; Role; Severities; Signal Pathway; Signal Transduction; Steroids; Structural defect; Structure; Structure of sinus venosus of fetus; Testing; Tretinoin; Vitamin A; Vitamins; apoAI regulatory protein-1; cardiogenesis; cofactor; congenital heart disorder; disease phenotype; functional genomics; genome-wide; human embryonic stem cell; improved; in vivo; induced pluripotent stem cell; insight; microCT; mouse model; novel; progenitor; recruit; response; stem cells; tool; transcription factor

ABSTRACT Deciphering the molecular circuitries that guide cardiac progenitor cell (CPC) differentiation will contribute to understand the causes of congenital heart disease (CHD). Vitamin A/Retinoic Acid (RA) signaling is a master regulator of cell specification in the heart. RA induces posterior specification of the cardiac progenitor cells (CPCs) of the Second Heart Field (pSHF), the pool that populates the atria and sinus venosus structures. Consequently, VitaminA/RA deficiency and excess in this developmental window lead to structural abnormalities in the heart. Thus, understanding the mechanisms that regulate RA signaling homeostasis is a major goal in the heart field. Here, we adopted a functional genomic approach at single-cell resolution to investigate the Retinoic Acid- regulatory network in human embryonic stem cell-derived CPCs (hESC-CPCs). Our preliminary data suggest that the Hippo effectors YAP1 and TEAD4 are non-canonical effectors of RA signaling critical for atrial cardiomyocyte differentiation. Our studies show that YAP deletion affects the expression of RA-target genes and phenotypic properties of atrial cardiomyocytes (CMs). Furthermore, our single-cell ATAC-seq analysis in WT versus YAP KO CPCs suggest that YAP facilitates accessibility of key atrial transcription factors (TFs) to the chromatin, including MEIS2, HOXA1 and GATA6, supporting a pivotal role of YAP in the TF composition of atrial enhancers (Aim1). Interestingly, our ongoing studies show that RA signaling induces genome-wide de-novo recruitment of YAP:TEAD4 to atrial enhancers in CPCs, independently of the activity of the Hippo-kinases. Our findings suggest that RA recruits YAP to the chromatin through the RA-activated steroid nuclear receptor NR2F2. NR2F2 is a key RA-effector, that regulates the acquisition of atrial lineages in hESC-CPCs and pSHF progenitors. Moreover human NR2F2 mutations lead to CHD. Therefore, we will investigate a novel non-canonical function of YAP and TEAD4 as NR2F2 cofactors in regulating atrial specification, highlighting the potential of YAP/TEAD as possible CHD modifiers (Aim2). Accordingly, our ongoing studies in mouse models and iPSCs revealed that the 22q11.2 genetic deletion or DiGeorge Syndrome (DGS) lead to the inactivation of YAP signaling. DGS is one of the main causes of human CHD, which is largely attributed to the haploinsufficiency in TBX1 in concert with downstream pathways, such as Vitamin A/RA. It has been shown that TBX1 loss in the SHF lead to unbalanced Vitamin A signaling, which evokes in erratic specification of these cells and CHD. Therefore, YAP inactivation in DGS may contribute to abnormal RA signaling in SHF cells and yet, CHD. Thus, we will examine whether restoring the activity of YAP in a mouse model of DGS improves CHD phenotypes. Finally, in collaboration with the UCLA CHD Biocore, we will validate the impact of the Retinoic Acid and YAP pathways in the phenotypes of biopsies of infants with CHD and DGS (Aim3). Findings from these aims will provide novel mechanist insights of normal CPC differentiation and into alterations that cause defective CPC differentiation in CHD.

抽象的 解密指导心脏祖细胞（CPC）分化的分子电路将有助于 了解先天性心脏病（CHD）的原因。维生素A/视黄酸（RA）信号是主人 心脏中细胞规范的调节剂。 RA诱导心脏祖细胞的后验规格 （CPC）的第二心脏场（PSHF），该池填充了心房和窦结构。 因此，这种发育窗口中的维生素/ra缺乏症和过量导致结构 心脏异常。因此，了解调节RA信号稳态的机制是一种 心脏领域的主要目标。在这里，我们采用了单细胞分辨率的功能基因组方法 研究人类干细胞衍生的CPC（HESC-CPC）中的视黄酸调节网络。 我们的初步数据表明，河马效应子yap1和tead4是RA的非典型效应子 信号对于心房心肌细胞分化至关重要。我们的研究表明，YAP缺失会影响 心房心肌细胞（CMS）的Ra-target基因和表型特性的表达。此外，我们的 WT与YAP KO CPC中的单细胞ATAC-SEQ分析表明，YAP促进了密钥的可访问性 染色质的心房转录因子（TFS），包括Meis2，Hoxa1和Gata6，支持关键 YAP在心房增强子的TF组成中的作用（AIM1）。有趣的是，我们正在进行的研究表明RA 信号传导可诱导全基因组的De-Novo募集YAP：TEAD4至CPC中心房增强子的募集， 独立于河马激酶的活性。我们的发现表明RA将YAP招募到染色质 通过RA激活的类固醇核受体NR2F2。 NR2F2是关键的RA效应器，可调节 在HESC-CPC和PSHF祖细胞中获取房屋谱系。此外，人NR2F2突变导致 冠心。因此，我们将研究YAP和TEAD4作为NR2F2辅因子的新型非传统功能 调节心房规范，突出显示YAP/TEAD作为可能的CHD修饰符的潜力（AIM2）。 因此，我们在小鼠模型和IPSC中正在进行的研究表明，22q11.2遗传缺失或 Digeorge综合征（DGS）导致YAP信号传导失活。 DGS是人类的主要原因之一 CHD主要归因于TBX1与下游途径的一致性归因 作为维生素A/RA。已经显示，SHF中的TBX1损失导致维生素A信号不平衡，该信号传导 这些细胞和冠心病的规格不稳定。因此，DG中的YAP失活可能有助于 SHF细胞中的RA信号异常，但CHD。因此，我们将检查是否恢复YAP的活动 在小鼠模型中，DGS改善了CHD表型。最后，在与UCLA CHD BIOCORE合作，我们 将验证视黄酸和YAP途径在婴儿活检表型中的影响 CHD和DGS（AIM3）。这些目标的发现将为正常CPC提供新颖的机械师见解 分化和变化导致CPC分化有缺陷的CHD。