Novel peptides for deciphering structural and signalling functions of E-cadherin in mouse embryonic stem cells.
Novel peptides for deciphering structural and signalling functions of E-cadherin in mouse embryonic stem cells.
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DOI：
10.1038/srep41827
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发表时间：
2017-02-07
影响因子：
4.6
通讯作者：
Ward CM
中科院分区：
文献类型：
--
作者：
Segal JM;Ward CM
我们之前已经表明，E-钙粘蛋白调节小鼠胚胎干细胞（mESC）的幼稚多能性状态，使LIF依赖性STAT 3磷酸化，与E-钙粘蛋白空mESC表现出超过3000个基因转录改变和激活素/Nodal依赖性多能性的开关。然而，阐明的确切机制与E-钙粘蛋白在胚胎干细胞的功能是复杂的，在描绘这种蛋白质的结构和信号功能的困难。在这里，我们表明，用E-钙粘蛋白中和抗体DECMA-1或E-钙粘蛋白结合肽H-SWELYYPLRANL-NH 2（Epep）处理的mESC表现出多能转录物和NANOG蛋白表达的离散谱，表明所用的E-钙粘蛋白抑制剂的类型决定了mESC的细胞表型。Epep的丙氨酸扫描突变揭示了Tbx 3、Klf 4和Esrrb转录抑制、细胞-细胞接触消除、细胞悬浮存活、STAT 3磷酸化和水溶性的关键残基。发现STAT 3磷酸化不依赖于细胞-细胞接触和激活素/Nodal依赖性多能性的丧失，并且描述了增强mESC中STAT 3磷酸化和Nanog转录物和蛋白质表达的肽。这些肽代表了一个有用的资源，用于破译E-钙粘蛋白的结构和信号传导功能，并证明完全缺乏E-钙粘蛋白蛋白可能需要分层的信号传导通路的改变mESC。
We have previously shown that E-cadherin regulates the naive pluripotent state of mouse embryonic stem cells (mESCs) by enabling LIF-dependent STAT3 phosphorylation, with E-cadherin null mESCs exhibiting over 3000 gene transcript alterations and a switch to Activin/Nodal-dependent pluripotency. However, elucidation of the exact mechanisms associated with E-cadherin function in mESCs is compounded by the difficulty in delineating the structural and signalling functions of this protein. Here we show that mESCs treated with the E-cadherin neutralising antibody DECMA-1 or the E-cadherin binding peptide H-SWELYYPLRANL-NH2 (Epep) exhibit discrete profiles for pluripotent transcripts and NANOG protein expression, demonstrating that the type of E-cadherin inhibitor employed dictates the cellular phenotype of mESCs. Alanine scanning mutation of Epep revealed residues critical for Tbx3, Klf4 and Esrrb transcript repression, cell-cell contact abrogation, cell survival in suspension, STAT3 phosphorylation and water solubility. STAT3 phosphorylation was found to be independent of loss of cell-cell contact and Activin/Nodal-dependent pluripotency and a peptide is described that enhances STAT3 phosphorylation and Nanog transcript and protein expression in mESCs. These peptides represent a useful resource for deciphering the structural and signalling functions of E-cadherin and demonstrate that complete absence of E-cadherin protein is likely required for hierarchical signalling pathway alterations in mESCs.