Department of Cell and Developmental Biology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, United States of America.
Institute for Biomedical Informatics, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, United States of America.
PLoS Biol. 2021 Jan 22;19(1):e3001059. doi: 10.1371/journal.pbio.3001059. eCollection 2021 Jan.
Bone Morphogenetic Protein (BMP) patterns the dorsal-ventral (DV) embryonic axis in all vertebrates, but it is unknown how cells along the DV axis interpret and translate the gradient of BMP signaling into differential gene activation that will give rise to distinct cell fates. To determine the mechanism of BMP morphogen interpretation in the zebrafish gastrula, we identified 57 genes that are directly activated by BMP signaling. By using Seurat analysis of single-cell RNA sequencing (scRNA-seq) data, we found that these genes are expressed in at least 3 distinct DV domains of the embryo. We distinguished between 3 models of BMP signal interpretation in which cells activate distinct gene expression through interpretation of thresholds of (1) the BMP signaling gradient slope; (2) the BMP signal duration; or (3) the level of BMP signal activation. We tested these 3 models using quantitative measurements of phosphorylated Smad5 (pSmad5) and by examining the spatial relationship between BMP signaling and activation of different target genes at single-cell resolution across the embryo. We found that BMP signaling gradient slope or BMP exposure duration did not account for the differential target gene expression domains. Instead, we show that cells respond to 3 distinct levels of BMP signaling activity to activate and position target gene expression. Together, we demonstrate that distinct pSmad5 threshold levels activate spatially distinct target genes to pattern the DV axis.
骨形态发生蛋白（BMP）在所有脊椎动物中塑造胚胎的背腹（DV）轴，但尚不清楚沿 DV 轴的细胞如何解释和转化 BMP 信号的梯度，从而激活不同的基因，产生不同的细胞命运。为了确定 BMP 形态发生因子在斑马鱼原肠胚中的解释机制，我们鉴定了 57 个直接受 BMP 信号激活的基因。通过对单细胞 RNA 测序（scRNA-seq）数据的 Seurat 分析，我们发现这些基因在胚胎的至少 3 个不同的 DV 区域表达。我们区分了 3 种 BMP 信号解释模型，细胞通过解释（1）BMP 信号梯度斜率；（2）BMP 信号持续时间；或（3）BMP 信号激活水平来激活不同的基因表达。我们使用定量测量磷酸化 Smad5（pSmad5）以及通过在单细胞分辨率下检查 BMP 信号与不同靶基因激活之间的空间关系来检验这 3 种模型。我们发现 BMP 信号梯度斜率或 BMP 暴露持续时间并不能解释不同靶基因表达域。相反，我们表明细胞对 3 种不同水平的 BMP 信号活性作出反应，以激活和定位靶基因表达。总之，我们证明了不同的 pSmad5 阈值水平激活空间上不同的靶基因，从而塑造 DV 轴。
