Identification of the C-terminal activation domain of the NeuroD-related factor (NDRF)

NeuroD-related factor (NDRF) is a basic helix–loop–helix (bHLH) protein whose expression is restricted to the central nervous system, and is considered to be responsible for maintenance of differentiated neurons as well as neurogenesis. NDRF structurally resembles NeuroD in the bHLH region and can induce neurogenesis ectopically in ectodermal cells of the Xenopus embryo. In this study, we delineated the functional domains of NDRF. Using GAL4/NDRF fusion proteins, we identified the C-terminal activation domain (C-AD) in NDRF between amino acid positions 294 and 383. This region was highly homologous to one part of the activation domain of NeuroD. We further investigated the transactivational function of C-AD in the mouse type 1 inositol 1,4,5-trisphosphate receptor promoter, which has an NDRF site. Truncation of C-AD resulted in reduction of the activation function, whereas the DNA-binding specificity was not affected. These results suggest that C-AD has a stimulatory function in the mammalian nervous system.     

Structural feature of sorghum chloroplast psbA gene and regulation effects of its 5'-noncoding region*

Comparative analysis reveals remarkable homology between the sequences of both psbA gene nucleotides and the inferred amino acids of sorghum, a C_4 plant, and those of rice, a C_3 plant. The 5'-noncoding region of sorghum psbA gene contains the conservative promoter elements, "-35" element and "-10" element, like the prokaryote and the promoter element, TATA box, like the eukaryote. As compared with that of the rice, an extra sequence of 7 bp is found in the leader sequence of the mRNA in the former. Using an in vitro system, it has been demonstrated that protein factor exists in sorghum chloroplast protein extract which specifically binds to the 5'-noncoding region of psbA gene. Measurement of the expression of luciferase shows a 2—5 time greater reaction of the expression plasmids pALqs which contain leader region of sorghum psbA gene than that of the expression plasmids pALqr which contain leader region of rice psbA gene in E. coli.     

ERα与Sp1相互作用激活LRP16启动子转录活性

根据已报道的LRP1 6启动子序列 (2 6kb) ,采用PCR反应获得 6个启动子 5′删除突变体 ,分别插入pGL3 Basic载体 ,构建 6种 5′缺失报告基因表达载体 (pS1 ～pS6) .分别与ERα真核表达载体共转染MCF 7细胞 .用双荧光素酶报告系统测定荧光素酶活性 ,以明确LRP1 6基因上游启动子区域中的雌激素反应序列 .结果显示 ,pS1 ～pS6均有雌二醇反应性 .进而对pS5的 3′端缺失分析发现LRP1 6基因翻译起始位点上游 - 2 1 4至 - 2 5 1位置的序列具有雌激素应答 ,序列分析发现该片段序列中包含了一个供转录因子Sp1结合的GC富含位点和一个ERα反应元件的半位点 (1 2ERE Sp1 ) ,进一步突变分析显示这两个元件均为雌激素反应性所必需 .以含这两个顺式元件的序列 (- 2 5 3bp至 - 2 2 4bp)作为探针 ,超级迁移凝胶电泳试验结果表明了ERα和Sp1蛋白均可以和探针结合 .研究发现了雌激素上调LRP1 6基因表达的一个增强子元件— 1 2ERE Sp1 ,ERα和Sp1蛋白需要与DNA结合形成复合体 ,通过其相互作用激活转录