Hamazaki Takashi, Kehoe Sarah M, Nakano Toru, Terada Naohiro
Department of Pathology, University of Florida College of Medicine, P.O. Box 100275, Gainesville, FL 32610, USA.
Mol Cell Biol. 2006 Oct;26(20):7539-49. doi: 10.1128/MCB.00508-06. Epub 2006 Aug 14.
The homeobox gene Nanog is a key intrinsic determinant of self renewal in embryonic stem (ES) cells, and its repression leads ES cells to selectively differentiate into primitive endoderm. Although Nanog repression occurs at the outermost layer of ES cell aggregates independent of the leukemia inhibitory factor (LIF)/STAT3 pathway, it is largely undetermined what external cues and intracellular signals cause the event. Of interest, addition of the tyrosine phosphatase inhibitor, sodium vanadate, selectively repressed Nanog transcription without any detectable changes in upstream transcriptional regulators Oct3/4 and Sox2. Furthermore, sodium vanadate induced primitive endoderm differentiation, even in the inner cells of ES cell aggregates. Expression of Gata6 and Zfp42, two putative downstream Nanog effectors, was also increased and decreased by the addition of sodium vanadate, respectively, but these changes were eliminated by exogenous Nanog expression. The effects of sodium vanadate were abrogated by Grb2 deficiency or by the addition of the Mek inhibitor, PD98059. Indeed, PD98059 prevented Nanog repression induced by ES cell aggregation as well. Furthermore, transfection of a constitutive active Mek mutant into ES cells induced Nanog repression and primitive endoderm differentiation. These data indicate that the Grb2/Mek pathway primarily mediates Nanog gene repression upon ES cell differentiation into primitive endoderm.
同源盒基因Nanog是胚胎干细胞自我更新的关键内在决定因素，其抑制会导致胚胎干细胞选择性分化为原始内胚层。尽管Nanog的抑制发生在胚胎干细胞聚集体的最外层，且独立于白血病抑制因子（LIF）/信号转导和转录激活因子3（STAT3）信号通路，但在很大程度上仍不清楚是什么外部信号和细胞内信号导致了这一事件。有趣的是，添加酪氨酸磷酸酶抑制剂钒酸钠可选择性地抑制Nanog转录，而上游转录调节因子八聚体结合转录因子3/4（Oct3/4）和性别决定区Y框蛋白2（Sox2）没有任何可检测到的变化。此外，即使在胚胎干细胞聚集体的内部细胞中，钒酸钠也能诱导原始内胚层分化。添加钒酸钠还分别增加和降低了两个假定的Nanog下游效应因子Gata6和锌指蛋白42（Zfp42）的表达，但这些变化可通过外源性Nanog表达消除。Grb2缺陷或添加丝裂原活化蛋白激酶激酶（Mek）抑制剂PD98059可消除钒酸钠的作用。事实上，PD98059也能阻止胚胎干细胞聚集诱导的Nanog抑制。此外，将组成型活性Mek突变体转染到胚胎干细胞中可诱导Nanog抑制和原始内胚层分化。这些数据表明，Grb2/Mek信号通路主要介导胚胎干细胞向原始内胚层分化时的Nanog基因抑制。
