Hwang Soon Baek, Lee Junho
National Research Laboratory, Department of Biology, Yonsei University, 134 Shinchon-dong, Seodaemun-ku, Seoul 120-749, South Korea.
J Mol Biol. 2003 Oct 17;333(2):237-47. doi: 10.1016/j.jmb.2003.08.055.
In order to characterize the mechanisms regulating neuronal expression of the nematode SNAP-25 gene, we identified the SNAP-25 genes of Caenorhabditis elegans and Caenorhabditis briggsae. Comparative sequence analysis and reporter assays revealed two putative 5' regulatory elements, P1 and P2, and four elements, I1h, I1m, I2h, and I2m, in the first intron. Nuclear extracts contained activities that bound the P2 and I1h elements. Different elements were required for SNAP-25 expression in different neuronal subsets; P1 was required in DA and DD motor neurons, and I1m and I2m were required in DB and DA neurons, respectively. P2 was active in amphid and phasmid neurons, I1h in pharyngeal neurons, and I2h in touch receptor neurons. The I2h element contained a putative binding site for transcription factor UNC-86. Both UNC-86 and MEC-3 were required for I2h activity in the mechanosensory neurons: in these neurons, GFP expression driven by I2h was abolished in animals bearing either an unc-86 null or a mec-3 null mutation, or an unc-86 mutation that leads to defective interaction with MEC-3. Deletion of the MEC-3 binding site also abolished the GFP expression. Gel mobility-shift assay results suggest that transcriptional regulation of SNAP-25 may involve multiple transcription factors.
为了阐明调控线虫SNAP - 25基因神经元表达的机制,我们鉴定了秀丽隐杆线虫和布里格隐杆线虫的SNAP - 25基因。比较序列分析和报告基因检测揭示了两个假定的5'调控元件P1和P2,以及位于第一个内含子中的四个元件I1h、I1m、I2h和I2m。核提取物含有与P2和I1h元件结合的活性。不同的神经元亚群中SNAP - 25表达需要不同的元件;DA和DD运动神经元中需要P1,DB和DA神经元中分别需要I1m和I2m。P2在化感器和尾感器神经元中具有活性,I1h在咽部神经元中具有活性,I2h在触觉感受器神经元中具有活性。I2h元件包含转录因子UNC - 86的一个假定结合位点。在机械感觉神经元中,I2h活性需要UNC - 86和MEC - 3两者:在这些神经元中,携带unc - 86无效突变、mec - 3无效突变或导致与MEC - 3相互作用缺陷的unc - 86突变的动物中,由I2h驱动的GFP表达被消除。MEC - 3结合位点的缺失也消除了GFP表达。凝胶迁移率变动分析结果表明,SNAP - 25的转录调控可能涉及多个转录因子。