Comprehensive identification and characterization of the binding sites on TLE in the Xenopus tropicalis gastrula.. Xenopus tropicalis

In vertebrates, homeodomain proteins Otx2 and Lim1/Lhx1 are required for head formation, but the regulatory principles underlying their functions in the head organizer remain unsolved. Here we show using ChIP-seq analysis that Otx2, Lim1, the coactivator p300 and the corepressor TLE/Groucho colocalize on cis-regulatory modules (CRMs) of thousands of genes including almost all ‘head-organizer’ genes in the Xenopus tropicalis gastrula. Comprehensive analysis of CRMs with RNA-seq data revealed that Lim1/Otx2-bound CRMs co-localizing with TLE rather than p300 are strongly associated with region/tissue-specific genes. Together with reporter analyses, our data suggest that Otx2 activates head-organizer genes with Lim1 and represses non-head-organizer genes with transcriptional repressors such as Goosecoid. Thus, it is likely that each of thousands of genes interprets Otx2 as a ‘positional tag’ to determine its expression in the head organizer, corroborating the idea that positional information directly contributes “massively parallel (distributive) gene regulation” rather than “hierarchal top-down gene control.” This general principle of gene regulation by region/tissue specific transcription factors may underlie the extraordinary diversities of morphologies in animals.

在脊椎动物中，同源域蛋白Otx2与Lim1/Lhx1是头部发生所必需的，但二者在头部组织者（head organizer）中发挥功能的调控机制仍未阐明。本研究通过染色质免疫共沉淀测序（ChIP-seq）分析证实，在热带爪蟾（Xenopus tropicalis）原肠胚中，Otx2、Lim1、共激活因子p300以及共抑制因子TLE/Groucho在数千个基因的顺式调控模块（CRMs）上共定位，其中几乎涵盖了所有‘头部组织者’相关基因。结合RNA测序（RNA-seq）数据对顺式调控模块进行综合分析后发现，与TLE共定位而非与p300共定位的Lim1/Otx2结合型顺式调控模块，与区域/组织特异性基因存在显著关联。结合报告基因分析结果，本研究数据表明：Otx2可与Lim1协同激活头部组织者相关基因，同时通过Goosecoid等转录抑制因子抑制非头部组织者基因的表达。由此推测，数千个基因均可将Otx2作为‘位置标签’，以此决定其在头部组织者中的表达模式，这一发现佐证了‘位置信息直接贡献于“大规模并行（分布式）基因调控”而非“层级式自上而下基因调控”’的观点。这种由区域/组织特异性转录因子介导的通用基因调控机制，可能是动物形态多样性产生的分子基础。