Kurokawa Daisuke, Kiyonari Hiroshi, Nakayama Rika, Kimura-Yoshida Chiharu, Matsuo Isao, Aizawa Shinichi
Laboratory for Vertebrate Body Plan, Center for Developmental Biology (CDB RIKEN Kobe, 2-2-3 Minatojima-minamimachi, Chuo-ku, Kobe 650-0046, Japan.
Development. 2004 Jul;131(14):3319-31. doi: 10.1242/dev.01220. Epub 2004 Jun 16.
Otx2 expression in the forebrain and midbrain was found to be regulated by two distinct enhancers (FM and FM2) located at 75 kb 5' upstream and 115 kb 3' downstream. The activities of these two enhancers were absent in anterior neuroectoderm earlier than E8.0; however, at E9.5 their regions of activity spanned the entire mesencephalon and diencephalon with their caudal limits at the boundary with the metencephalon or isthmus. In telencephalon, activities were found only in the dorsomedial aspect. Potential binding sites of OTX and TCF were essential to FM activity, and TCF sites were also essential to FM2 activity. The FM2 enhancer appears to be unique to rodent; however, the FM enhancer region is deeply conserved in gnathostomes. Studies of mutants lacking FM or FM2 enhancer demonstrated that these enhancers indeed regulate Otx2 expression in forebrain and midbrain. Development of mesencephalic and diencephalic regions was differentially regulated in a dose-dependent manner by the cooperation between Otx1 and Otx2 under FM and FM2 enhancers: the more caudal the structure the higher the OTX dose requirement. At E10.5 Otx1-/-Otx2DeltaFM/DeltaFM mutants, in which Otx2 expression under the FM2 enhancer remained, exhibited almost complete loss of the entire diencephalon and mesencephalon; the telencephalon did, however, develop.
研究发现，前脑和中脑中Otx2的表达受位于其5'上游75 kb和3'下游115 kb处的两个不同增强子（FM和FM2）调控。在早于E8.0的前神经外胚层中，这两个增强子均无活性；然而，在E9.5时，它们的活性区域跨越整个中脑和间脑，其尾端界限位于与后脑或峡部的边界处。在端脑中，活性仅见于背内侧。OTX和TCF的潜在结合位点对FM活性至关重要，TCF位点对FM2活性也至关重要。FM2增强子似乎是啮齿动物所特有的；然而，FM增强子区域在有颌类动物中高度保守。对缺乏FM或FM2增强子的突变体的研究表明，这些增强子确实调控前脑和中脑中Otx2的表达。在FM和FM2增强子的作用下，Otx1和Otx2的协同作用以剂量依赖的方式对中脑和间脑区域的发育进行差异调控：结构越靠尾端，OTX剂量需求越高。在E10.5的Otx1-/-Otx2DeltaFM/DeltaFM突变体中，FM2增强子下的Otx2表达得以保留，该突变体几乎完全缺失整个间脑和中脑；然而，端脑却发育了。
