Nakamura Hisashi, Tashiro Kosuke, Shiokawa Koichiro
Laboratory of Molecular Embryology, Zoological Institute, Faculty of Science, University of Tokyo, 7-3-1 Hongo, 113, Tokyo, Japan.
Rouxs Arch Dev Biol. 1996 Feb;205(5-6):300-310. doi: 10.1007/BF00365808.
Previously, we isolated Xenopus HGF (hepatocyte growth factor) cDNA and showed in Xenopus embryos that expression of this gene starts at the late gastrula stage mainly in the ventral mesoderm, and furthermore that the expression is induced in animal cap by activin A and bFGF (basic fibroblast growth factor). Here we have cloned the Xenopus HGF gene, covering a 14 kb 5'-upstream region and a 0.2 kb 5'-coding region. Within about 0.5 kb of the 5'-flanking region, the Xenopus HGF gene contained a TATA-like element AATGAAA, one putative NF-1 binding site, two NF-IL-6 binding motif sequences, one putative TGF-β-dependent inhibitory element (TIE) and one AP-1 binding site. A recombinant circular plasmid consisting of a 1.7 kb HGF promoter region and the bacterial chloramphenicol acetyltransferase (CAT) gene was first expressed at the late gastrula stage in the ventral mesoderm, as was the endogenous HGF gene. The expression of the fusion gene was induced in animal cap cells by activin A and bFGF although induction by the latter was not so strong. Using a series of 5'-deletion constructs introduced into animal caps, silencer elements, which seem to be essential for the gene's regionally correct expression, and the element responsible for induction by activin were found. The results show that the HGF gene promoter isolated here contains elements which may endow the gene with the regulative function for its temporally and spatially regulated expression, although the element necessary for induction by bFGF seems to be missing.
此前，我们分离出了非洲爪蟾肝细胞生长因子（HGF）的cDNA，并在非洲爪蟾胚胎中发现，该基因在原肠胚晚期开始表达，主要表达于腹侧中胚层。此外，在动物帽中，激活素A和碱性成纤维细胞生长因子（bFGF）可诱导该基因的表达。在此，我们克隆了非洲爪蟾HGF基因，其涵盖了一个14 kb的5'上游区域和一个0.2 kb的5'编码区域。在5'侧翼区域约0.5 kb范围内，非洲爪蟾HGF基因包含一个类TATA元件AATGAAA、一个假定的NF-1结合位点、两个NF-IL-6结合基序序列、一个假定的转化生长因子-β（TGF-β）依赖性抑制元件（TIE）和一个AP-1结合位点。一个由1.7 kb的HGF启动子区域和细菌氯霉素乙酰转移酶（CAT）基因组成的重组环状质粒，最初在原肠胚晚期于腹侧中胚层表达，内源性HGF基因也是如此。激活素A和bFGF可在动物帽细胞中诱导融合基因的表达，不过后者的诱导作用没那么强。通过将一系列5'缺失构建体导入动物帽，发现了沉默元件，这些元件似乎对该基因在区域上的正确表达至关重要，还发现了负责激活素诱导的元件。结果表明，此处分离出的HGF基因启动子包含一些元件，这些元件可能赋予该基因对其时空调节表达的调控功能，尽管bFGF诱导所必需的元件似乎缺失了。
