Analysis of HLH factors regulating vertebrate neurogenesis

在哺乳动物的神经发育过程中，各种各样的神经元从相对统一的普通前体细胞中分化出来。众所周知，细胞间相互作用是产生神经元类型多样性的必要条件；分化神经元抑制邻近细胞分化为相同的神经元类型（侧抑制）。虽然跨膜蛋白Notch参与了侧抑制过程，但人类Notch的细胞内通路仍然非常难以捉摸。最近，我们等人发现bHLH基因Hes1和Hes5的启动子活性是由Notch激活诱导的。因此，为了了解哺乳动物Notch通路，我们在野生型、Hes1-空型、Hes5-空型和Hes1-Hes5双空型小鼠胚胎制备的神经前体细胞中，通过逆转录病毒错误表达构成活性形式的Notch （caNotch）来检测Hes1和Hes5的作用。我们发现，caNotch诱导内源性Hes1和Hes5表达，在野生型、Hes1 -null和Hes5 -null背景下抑制神经元分化，而在Hes1 -Hes5双空背景下无抑制作用。这些结果表明Hes1和Hes5在哺乳动物神经元分化调控中是必不可少的Notch效应因子，并且这两个hes基因在Notch下游相互补偿。我们之前的研究表明，Hes通过拮抗bhlh型神经元决定/分化基因Mash1抑制神经元分化。综上所述，这些数据表明哺乳动物神经元分化受“Notch * Hes1/Hes5 * Mash1 *神经元分化”通路调控（*，激活；*，抑制）。

During mammalian neural development, a wide variety of neurons differentiates from relatively uniform common precursor cells. It is known that cell-cell interaction is essential to generate the neuronal type diversity ; Differentiating neurons inhibit the neighboring cells from differentiating into the same neuronal types (lateral inhibition). While the transmembrane protein Notch is involved in the lateral inhibition process, the intracellular pathway of manunalian Notch remains very elusive.Recently, we and others found that the promoter activity of the bHLH genes Hes1 and Hes5 was induced by activation of Notch. Thus, to understand the mammalian Notch pathway, the role of Hes1 and Hes5 was examined by retrovirally misexpressing the constitutively active form of Notch (caNotch) in neural precursor cells prepared from wild-type, Hes1-null, Hes5-null, and Hes1-Hes5 double-null mouse embryos. We found that caNotch, which induced the endogenous Hes1 and Hes5 expression, inhibited neuronal differentiation in the wild-type, Hes1 -null, and Hes5-nulI background but not in the Hes1 -Hes5 double-null background. These results demonstrate that Hes1 and Hes5 are essential Notch effectors in regulation of mammalian neuronal differentiation and that the two Hesgenes conpensate each other downstream of Notch. We previously showed that Hes] inhibits neuronal differentiation by antagonizing the bHLH-type neuronal determination/differentiation gene Mash1.Taken together, these data indicate that mammalian neuronal differentiation is regulated by the pathway "Notch * Hes1/Hes5 * Mash1 * neuronal differentiation" (*, activation ; * , inhibition).