A Retinoic Acid:YAP1 signaling axis controls atrial lineage commitment

Vitamin A/Retinoic Acid (RA) signaling is essential for heart development. In cardiac progenitor cells (CPCs), RA signaling induces the expression of atrial lineage genes while repressing ventricular genes, thereby promoting the acquisition of an atrial cardiomyocyte cell fate. To achieve this, RA coordinates a complex regulatory network of downstream effectors that is not fully identified. To address this gap, we applied a functional genomics approach (scRNAseq, scATACseq and ChIP-seq) to untreated and RA-treated human embryonic stem cells (hESCs)-derived CPCs. Unbiased analysis revealed that the Hippo effectors YAP and TEAD4 are integrated with the atrial transcription factor enhancer network, and that YAP1 is necessary for activation of RA-enhancers in CPCs. Furthermore, scRNAseq analysis of control and conditionally YAP KO mouse E7.75 embryos (Sox2cre) revealed that the expression of atrial lineage genes such as NR2F2 is compromised by YAP deletion in the CPCs of the second heart field. Accordingly, we found that YAP is required for the formation of an atrial chamber but is dispensable for the formation of a ventricle, in hESC-derived patterned cardiac organoids. Overall, our findings revealed that YAP1 is a non-canonical effector of RA signaling essential for the acquisition of atrial lineages during cardiogenesis. Single-cell RNA sequencing (scRNAseq) data were obtained from E7.75 mouse embryos, including cYAP KO (Sox2cre:Yapflox/flox) and control (Sox2cre:Yapflox/+).视黄酸（RA）信号通路对心脏发育至关重要。在心脏祖细胞（CPCs）中，RA信号可诱导心房谱系基因的表达，同时抑制心室基因的转录，进而促进细胞向心房心肌细胞命运分化。为解析这一过程的调控机制，RA会调控一套尚未完全明确的下游效应因子复杂调控网络。为填补这一研究空白，我们对未经RA处理及经RA处理的人类胚胎干细胞（hESCs）来源的心脏祖细胞，应用了功能基因组学研究策略，涵盖单细胞RNA测序（scRNAseq）、单细胞转座酶可及性测序（scATACseq）与染色质免疫共沉淀测序（ChIP-seq）。无偏分析结果显示，Hippo通路效应因子YAP与TEAD4可整合进入心房转录因子增强子调控网络，且YAP1对于心脏祖细胞中RA应答增强子的激活不可或缺。此外，我们对条件性YAP敲除的E7.75小鼠胚胎（Sox2cre）及其对照胚胎开展scRNAseq分析，发现第二心区心脏祖细胞中YAP的缺失会导致NR2F2等心房谱系基因的表达受损。相应地，我们在人类胚胎干细胞来源的模式化心脏类器官中发现，YAP是心房腔形成所必需的，但对心室腔的形成并非必需。综上，本研究揭示YAP1作为RA信号通路的非经典效应因子，在心脏发生过程中心房谱系的获得中发挥关键作用。本研究的单细胞RNA测序（scRNAseq）数据来源于E7.75阶段的小鼠胚胎，样本包括cYAP敲除组（Sox2cre:Yapflox/flox）与对照组（Sox2cre:Yapflox/+）。