A study on expression of the gene encoding photolyase for cyclobutane pyrimidine dimer and regulation mechanism of photolyase activity

UV-B对黄瓜初叶生长有抑制作用。DNA损伤光恢复酶对生长迟缓的作用谱与光对DNA损伤的作用谱相似，表明该酶与植物的紫外耐受性有关。因此，我研究阐明一种光恢复酶（光解酶）基因表达的调控系统。我从黄瓜植株中分离到了编码cpd -光解酶的cDNA和基因组DNA (CsPHR)， CsPHR的转录本（mRNA）在光周期的早晨达到最大值。由于UV-B的加入，今早的峰值变高了。我做了光诱导CsPHR的作用谱，光谱显示在310nm左右诱导的峰值。接下来，我将嵌合体基因GUS基因作为与CsPHR启动子结合的报告基因引入拟南芥。在含UV-B的光照射下，GUS基因在叶尖、茎和根中表达。在本案例中，GUS诱导作用谱中最显著的诱导峰位于310nm左右。将含有报告基因LUC的启动子的合成衍生物引入拟南芥中，明确了启动子的顺式结构域，并研究了UV-B对LUC的诱导作用。结果发现，在翻译起始点上游202-296bp的一个启动子域是诱导所必需的顺式结构域。然而，也表明该域不可能单独诱导报告子，提示在翻译起始上游201bp内存在其他因素与该域协同作用。

Growth of cucumber first leaves was retarded by UV-B. It was suggested that light recovery enzyme of DNA damage was concerned with UV tolerance of plants, because the action spectrum for growth retardation was similar to the spectrum for DNA damage caused by light. Thus, I studied to elucidate control system of gene expression of a light recovery enzyme (photolyase). I isolated the cDNA, which encoded CPD-photolyase, and the genome DNA (CsPHR) from cucumber plants The level of transcript (mRNA) of CsPHR reached a maximum in the morning in the photoperiod. The peak of this morning became higher by UV-B addition.I made the action spectrum of CsPHR induction by light, and the spectrum showed the highest peak of induction around 310nm. Next, I introduced the chimera gene, GUS gene as a reporter bound to the promoter of CsPHR, into Arabidopsis. GUS was expressed in the tip of leaves, the stem and the roots when light including UV-B was irradiated to these transgenic plants. In this case too, the most prominent peak of the induction was observed around 310nm in the action spectrum for GUS induction.Synthetic derivatives of the promoter harboring a reporter gene LUC were introduced into Arabidopsis, to clarify cis domain of the promoter, and the induction of LUC by UV-B was examined. It was found that a promoter domain of 202-296bp upper stream of the translation start point was a cis domain essential for the induction. However, it was also showed that this domain alone was not possible to induce the reporter, and it was suggested that there are other factors which work cooperatively with the domain within 201bp upper stream of the translation start.