Department of Biological Sciences, Graduate School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Development. 2012 May;139(9):1651-61. doi: 10.1242/dev.068395.
How multiple developmental cues are integrated on cis-regulatory modules (CRMs) for cell fate decisions remains uncertain. The Spemann-Mangold organizer in Xenopus embryos expresses the transcription factors Lim1/Lhx1, Otx2, Mix1, Siamois (Sia) and VegT. Reporter analyses using sperm nuclear transplantation and DNA injection showed that cerberus (cer) and goosecoid (gsc) are activated by the aforementioned transcription factors through CRMs conserved between X. laevis and X. tropicalis. ChIP-qPCR analysis for the five transcription factors revealed that cer and gsc CRMs are initially bound by both Sia and VegT at the late blastula stage, and subsequently bound by all five factors at the gastrula stage. At the neurula stage, only binding of Lim1 and Otx2 to the gsc CRM, among others, persists, which corresponds to their co-expression in the prechordal plate. Based on these data, together with detailed expression pattern analysis, we propose a new model of stepwise formation of the organizer, in which (1) maternal VegT and Wnt-induced Sia first bind to CRMs at the blastula stage; then (2) Nodal-inducible Lim1, Otx2, Mix1 and zygotic VegT are bound to CRMs in the dorsal endodermal and mesodermal regions where all these genes are co-expressed; and (3) these two regions are combined at the gastrula stage to form the organizer. Thus, the in vivo dynamics of multiple transcription factors highlight their roles in the initiation and maintenance of gene expression, and also reveal the stepwise integration of maternal, Nodal and Wnt signaling on CRMs of organizer genes to generate the organizer.
多种发育线索如何在顺式调控模块（CRMs）上整合以做出细胞命运决定尚不清楚。非洲爪蟾胚胎的 Spemann-Mangold 组织者表达转录因子 Lim1/Lhx1、Otx2、Mix1、Siamois（Sia）和 VegT。使用精子核移植和 DNA 注射进行的报告基因分析表明，cerberus（cer）和 goosecoid（gsc）通过非洲爪蟾和热带爪蟾之间保守的 CRM 被上述转录因子激活。对这五个转录因子进行的 ChIP-qPCR 分析表明，cer 和 gsc CRM 在晚期囊胚期最初被 Sia 和 VegT 结合，随后在原肠胚期被所有五个因子结合。在神经胚期，只有 Lim1 和 Otx2 与 gsc CRM 的结合持续存在，这与它们在脑前板中的共表达相对应。基于这些数据，结合详细的表达模式分析，我们提出了组织者逐步形成的新模型，其中：（1）母体 VegT 和 Wnt 诱导的 Sia 首先在囊胚期结合 CRM；然后（2）Nodal 诱导的 Lim1、Otx2、Mix1 和合子 VegT 结合到在这些基因共表达的背内胚层和中胚层区域的 CRM；和（3）这两个区域在原肠胚期结合形成组织者。因此，多种转录因子的体内动力学突出了它们在基因表达的启动和维持中的作用，也揭示了母体、Nodal 和 Wnt 信号在组织者基因的 CRM 上的逐步整合，以产生组织者。
