Genome-wide Analysis of Smad7-Mediated Transcription in Mouse Embryonic Stem Cells [ChIP-Seq]

Smad7 has been identified as a negative regulator of the transforming growth factor TGF-ß pathway by direct interaction with the TGF-ß type I receptor (TßR-I). Although Smad7 has also been shown to play TGF-ß unrelated functions in the cytoplasm and in the nucleus, a comprehensive analysis of its nuclear function has not yet been performed. Here we show that in ESCs Smad7 is mainly nuclear and acts as a general transcription factor regulating a number of genes unrelated to the TGF-ß pathway. Loss of Smad7 results in the downregulation of several key stemness master regulators, including Pou5f1 and Zfp42, and in the upregulation of developmental genes, with consequent loss of the stem phenotype. Integrative analysis of genome-wide mapping data for Smad7 and ESC self-renewal and pluripotency transcriptional regulators revealed that Smad7 co-occupies promoters of highly expressed key stemness regulators genes, by binding to a specific consensus response element NCGGAAMM. Altogether, our data establishes Smad7 as new integral component of the regulatory circuitry that controls ESC identity. Overall design: ChIP-seq profiling of Bio-Smad7 in mouse Embryonic Stem Cells (ESC). 3 samples, 2 ChIP and 1 Mock as control.

Smad7已被证实可通过与转化生长因子β（transforming growth factor β, TGF-β）I型受体（TβR-I）直接相互作用，作为TGF-β通路的负调控因子。尽管已有研究表明Smad7在细胞质与细胞核中可发挥不依赖于TGF-β的功能，但目前尚未有针对其核功能的系统性全面分析。本研究证实，在小鼠胚胎干细胞（Embryonic Stem Cells, ESCs）中，Smad7主要定位于细胞核，并作为通用转录因子调控一系列与TGF-β通路无关的基因。Smad7的缺失会导致包括Pou5f1与Zfp42在内的多个关键干性主调控因子表达下调，同时使发育相关基因表达上调，最终引发干细胞干性表型的丧失。通过对Smad7全基因组结合图谱数据与ESC自我更新及多能性转录调控因子的整合分析发现，Smad7可通过结合特定保守应答元件NCGGAAMM，共同占据高表达的关键干性调控基因的启动子区域。综上，本研究数据确立Smad7为调控ESC身份的调控环路的全新核心组成部分。实验整体设计：对小鼠胚胎干细胞中的Bio-Smad7进行染色质免疫沉淀测序（ChIP-seq）分析。共设置3个样本，其中2个为ChIP实验组，1个为Mock对照样本。