Bulger Emily A, McDevitt Todd C, Bruneau Benoit G
Gladstone Institutes, San Francisco, CA.
Developmental and Stem Cell Biology Graduate Program, University of California, San Francisco, CA.
bioRxiv. 2024 Jan 26:2024.01.25.577277. doi: 10.1101/2024.01.25.577277.
Proper regulation of gene dosage is critical for the development of the early embryo and the extraembryonic tissues that support it. Specifically, loss of leads to stunted development of the allantois, an extraembryonic mesoderm-derived structure critical for nutrient delivery and waste removal in the early embryo. In this study, we investigate how CDX2 dose-dependently influences the gene regulatory network underlying extraembryonic mesoderm development. We generate an allelic series for in human induced pluripotent stem cells (hiPSCs) consisting of WT, heterozygous, and homozygous null genotypes, differentiate these cells in a 2D gastruloid model, and subject these cells to multiomic single nucleus RNA and ATAC sequencing. We identify several genes that CDX2 dose-dependently regulate cytoskeletal integrity and adhesiveness in the extraembryonic mesoderm population, including regulators of the VEGF, canonical WNT, and non-canonical WNT signaling pathways. Despite these dose-dependent gene expression patterns, snATAC-seq reveals that heterozygous CDX2 expression is capable of inducing a WT-like chromatin accessibility profile, suggesting accessibility is not sufficient to drive gene expression when the CDX2 dosage is reduced. Finally, because the loss of CDX2 or TBXT phenocopy one another , we compare differentially expressed genes in our CDX2 knock-out model to those from TBXT knock-out hiPSCs differentiated in an analogous experiment. This comparison identifies several communally misregulated genes that are critical for cytoskeletal integrity and tissue permeability, including and . Together, these results clarify how CDX2 dose-dependently regulates gene expression in the extraembryonic mesoderm and suggest these genes may underlie the defects in vascular development and allantoic elongation seen in the absence or reduction of CDX2 .
基因剂量的适当调控对于早期胚胎及其支持的胚外组织的发育至关重要。具体而言，[基因名称]的缺失会导致尿囊发育迟缓，尿囊是一种胚外中胚层衍生结构，对早期胚胎的营养输送和废物清除至关重要。在本研究中，我们研究了CDX2如何剂量依赖性地影响胚外中胚层发育的基因调控网络。我们在人类诱导多能干细胞（hiPSC）中生成了一个[基因名称]的等位基因系列，包括野生型、杂合子和纯合子缺失基因型，在二维类原肠胚模型中分化这些细胞，并对这些细胞进行多组学单核RNA和ATAC测序。我们鉴定了几个基因，CDX2以剂量依赖性方式调节胚外中胚层群体中的细胞骨架完整性和粘附性，包括VEGF、经典WNT和非经典WNT信号通路的调节因子。尽管有这些剂量依赖性基因表达模式，snATAC-seq显示杂合子CDX2表达能够诱导类似野生型的染色质可及性图谱，表明当CDX2剂量降低时，可及性不足以驱动基因表达。最后，由于CDX2或TBXT的缺失表现相似，我们将CDX2敲除模型中差异表达的基因与在类似实验中分化的TBXT敲除hiPSC中的基因进行比较。这种比较确定了几个共同失调的基因，这些基因对细胞骨架完整性和组织通透性至关重要，包括[基因名称1]和[基因名称2]。总之，这些结果阐明了CDX2如何剂量依赖性地调节胚外中胚层中的基因表达，并表明这些基因可能是在CDX2缺失或减少时所见的血管发育和尿囊伸长缺陷的基础。
