Tbx20 Is Required in Mid-Gestation Cardiomyocytes and Plays a Central Role in Atrial Development.
Tbx20 Is Required in Mid-Gestation Cardiomyocytes and Plays a Central Role in Atrial Development.
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DOI：
10.1161/circresaha.118.311339
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发表时间：
2018-08-03
影响因子：
20.1
通讯作者：
Evans SM
中科院分区：
文献类型：
--
作者：
Boogerd CJ;Zhu X;Aneas I;Sakabe N;Zhang L;Sobreira DR;Montefiori L;Bogomolovas J;Joslin AC;Zhou B;Chen J;Nobrega MA;Evans SM
补充数字内容可在正文中找到。转录因子TBX20(T-box 20)的突变与先天性心脏病有关。胚系消融Tbx20会导致心脏发育异常和胚胎死亡。由于Tbx20在心肌发育所需的多种细胞系中表达，包括咽内胚层、心源性中胚层、心内膜和心肌，因此在心肌发育早期对Tbx20的细胞类型特异性需求仍有待探索。在这里，我们研究了TBX20在妊娠中期心肌细胞在心脏发育中的作用。用多西环素诱导的cTnTCre转基因去除发育中的心肌细胞中的Tbx20会导致胚胎死亡。在Tbx20条件基因敲除突变体中，发育中的室腔和房室的周长，特别是未来左心房的周长显著减少。细胞周期分析显示，Tbx20突变的心肌细胞增殖减少，并停滞在G1-S期。对突变的心肌细胞的全基因组转录组分析显示，对细胞周期调节至关重要的多个基因的差异表达。此外，房室基因程序似乎受到了异常的调控。用染色质免疫沉淀高通量测序技术从胚胎心脏中鉴定了可能的直接靶点，包括关键的细胞周期基因和房室特异基因。值得注意的是，TBX20结合了一个对心房发育和识别至关重要的基因的保守增强子，Coup-TFII/Nr2f2(鸡卵蛋白上游启动子转录因子2/核受体亚家族2，F组，成员2)。这种增强子与人心肌细胞中的NR2F2启动子相互作用，并以TBX20依赖的方式在转基因小鼠中实现心房特异性基因的表达。心肌Tbx20直接调节胎儿心肌细胞增殖所需的一组基因，包括那些G1-S转换所需的基因。TBX20还直接下调祖细胞特异性基因，除了调节腔室心肌与非腔室心肌的基因外，还直接激活建立或维持心房和心室特性所需的基因。TBX20通过直接调节进化上保守的COUPT-FII增强子，在心房发育中发挥着以前未被认识的关键作用。
Supplemental Digital Content is available in the text. Mutations in the transcription factor TBX20 (T-box 20) are associated with congenital heart disease. Germline ablation of Tbx20 results in abnormal heart development and embryonic lethality by embryonic day 9.5. Because Tbx20 is expressed in multiple cell lineages required for myocardial development, including pharyngeal endoderm, cardiogenic mesoderm, endocardium, and myocardium, the cell type–specific requirement for TBX20 in early myocardial development remains to be explored. Here, we investigated roles of TBX20 in midgestation cardiomyocytes for heart development. Ablation of Tbx20 from developing cardiomyocytes using a doxycycline inducible cTnTCre transgene led to embryonic lethality. The circumference of developing ventricular and atrial chambers, and in particular that of prospective left atrium, was significantly reduced in Tbx20 conditional knockout mutants. Cell cycle analysis demonstrated reduced proliferation of Tbx20 mutant cardiomyocytes and their arrest at the G1-S phase transition. Genome-wide transcriptome analysis of mutant cardiomyocytes revealed differential expression of multiple genes critical for cell cycle regulation. Moreover, atrial and ventricular gene programs seemed to be aberrantly regulated. Putative direct TBX20 targets were identified using TBX20 ChIP-Seq (chromatin immunoprecipitation with high throughput sequencing) from embryonic heart and included key cell cycle genes and atrial and ventricular specific genes. Notably, TBX20 bound a conserved enhancer for a gene key to atrial development and identity, COUP-TFII/Nr2f2 (chicken ovalbumin upstream promoter transcription factor 2/nuclear receptor subfamily 2, group F, member 2). This enhancer interacted with the NR2F2 promoter in human cardiomyocytes and conferred atrial specific gene expression in a transgenic mouse in a TBX20-dependent manner. Myocardial TBX20 directly regulates a subset of genes required for fetal cardiomyocyte proliferation, including those required for the G1-S transition. TBX20 also directly downregulates progenitor-specific genes and, in addition to regulating genes that specify chamber versus nonchamber myocardium, directly activates genes required for establishment or maintenance of atrial and ventricular identity. TBX20 plays a previously unappreciated key role in atrial development through direct regulation of an evolutionarily conserved COUPT-FII enhancer.