Saitsu Hirotomo, Komada Munekazu, Suzuki Misao, Nakayama Rika, Motoyama Jun, Shiota Kohei, Ishibashi Makoto
Department of Anatomy and Developmental Biology, Graduate School of Medicine, Kyoto University, Yoshida, Sakyo-ku, Kyoto 606-8501, Japan.
Dev Dyn. 2005 Feb;232(2):282-92. doi: 10.1002/dvdy.20236.
Sonic hedgehog (Shh) is a secreted molecule that is thought to regulate tissue growth and patterning in vertebrate embryos. Although it has been reported that Gli transcription factors mediate Shh signaling to the nucleus, little is known about developmental target genes of Gli. In the previous genetic study, we showed that Shh is required for Fgf15 expression in the diencephalon and midbrain. Here, we examined whether Fgf15 is a direct target of Shh signaling through Gli. Shh was expressed in the midline cells and Fgf15 in the medial region of the diencephalon/midbrain by the seven-somite stage. The Fgf15 expression domain coincided with that of Gli1 and overlapped with that of Gli2 at this stage. Fgf15 expression in the diencephalon/midbrain was greatly reduced in the seven-somite Shh mutant embryos. Transgenic analysis showed that the 3.6-kb 5'-flanking region of the Fgf15 gene is sufficient for induction of Fgf15 in the medial/ventral diencephalon/midbrain. Luciferase assay showed that the 3.6-kb Fgf15 enhancer/promoter was activated by Gli2. A Gli-binding site was located 1 kb upstream of the transcription start site and was required for expression in the medial/ventral diencephalon/midbrain in transgenic embryos and for activation in luciferase assay. These findings indicate that Fgf15 is directly regulated by Shh signaling through Gli proteins.
音猬因子（Shh）是一种分泌型分子，被认为在脊椎动物胚胎中调节组织生长和模式形成。尽管已有报道称Gli转录因子介导Shh信号传导至细胞核，但对于Gli的发育靶基因知之甚少。在之前的遗传学研究中，我们发现Shh是间脑和中脑中Fgf15表达所必需的。在此，我们研究了Fgf15是否是Shh信号通过Gli的直接靶标。在七体节期，Shh在中线细胞中表达，Fgf15在间脑/中脑的内侧区域表达。此时，Fgf15的表达域与Gli1的表达域重合，并与Gli2的表达域重叠。在七体节Shh突变胚胎中，间脑/中脑中Fgf15的表达大幅降低。转基因分析表明，Fgf15基因的3.6 kb 5'侧翼区域足以在间脑/中脑的内侧/腹侧诱导Fgf15的表达。荧光素酶测定表明，3.6 kb的Fgf15增强子/启动子被Gli2激活。一个Gli结合位点位于转录起始位点上游1 kb处，是转基因胚胎中间脑/中脑内侧/腹侧表达以及荧光素酶测定中激活所必需的。这些发现表明，Fgf15是通过Gli蛋白由Shh信号直接调控的。
