Biochemical characteristics of cyclobutane pyrimidine dimer photolyase and UV-B protecting mechanisms of mice (Oryza sativa L.)

在2000年和2002.1年间，我们有了几个新的发现：对紫外线敏感的NICE Norin 1缺乏对环丁烷嘧啶食用者(CPDS)的光修复。利用闪光灯的方法，我们发现体内光修复的缺陷是由于结构/功能改变的光解酶所致。结果表明，Norin-1光解酶-二聚体复合体相对于耐紫外线的Sasanishiki光解酶是高度不耐热的。2.克隆了Sasanishiki和Norin-1的CPD光解酶基因。它们被发现编码506个氨基酸残基的ORF。Sasanishiki的第126位氨基酸是谷氨酰胺，Norin 1.3：GST融合的水稻CPD光解酶是精氨酸。我们从大肠杆菌KY20中纯化了CPD光解酶，确定FAD是两个发色团之一。4：暗生长的黄化水稻幼苗几乎没有CPD光解酶活性。暗生长的水稻幼苗中CPD光解酶的诱导是在光敏色素的光控制下进行的，而光解酶基因的表达是在光敏色素和/或隐花色素的光控制下进行的。5：我们利用农杆菌转化系统获得了转CPD光解酶基因的转基因水稻植株。6：尽管紫外光吸收化合物和花青素的积累高于T-65，但遗传背景为台中65的紫叶基因p1(T-65)的水稻近等基因系(NIL)的生长在UV-B升高的培养过程中显著低于T-65。NIL的稳态CPD水平也高于T-65。我们认为，在可见光照射下的生长受阻和CPD水平的增加可能是由于UV-B对产生CPD水平有效的UV-B和对CPD光解酶活性有效的蓝/UV-A的筛选效果不同所致。

We made several new findings in the period of 2000 and 2002.1:UV-sensitive nice Norin 1 is deficient in photorepair of cyclobutane pyrimidine diners (CPDs). Using a photoflash approach, we showed that the deficiency in photorepair in vivo resulted from a structure/function alteration photolyase. These results were confirmed by studies with extracts, which showed that the Norin 1 photolyase-dimer complex was highly thermolabile relative to the UV-resistant Sasanishiki photolyase.2:CPD photolyase gene from Sasanishiki and Norin 1 were cloned and sequenced. They were found to encode an ORF for 506 amino acid residues. Amino acid at position 126 was glutamine in Sasanishiki, but was arginine in Norin 1.3:GST-fusion rice CPD photolyase was purified from E. coli KY20, and we determined FAD as one of the two chromophores.4:Dark-grown etiolated rice seedlings did hardly have CPD photolyase activity. The induction of CPD photolyase in dark-grown seedlings was under photocontrol of phytochrome, and the expression of photolyase gene was under photocontrol of phytochrome and/or cryptochrome.5:We started to produce a transgenic rice plant into which CPD photolyase gene was transformed using the Agrobacterium transformation system.6:The growth of near-isogenic line (NIL) of rice for purple leaf gene pl of rice with a genetic background of Taichung 65 (T-65) was significantly decreased relative to T-65 in the elevated UV-B during culture regardless of higher accumulation of UV-absorbing compounds and anthocyanins than T-65. The steady-state CPD levels in the NIL was also higher than that in T-65. We showed that the growth stunting and increase in CPD levels under chronic UV-B irradiated with visible light might be due to the difference in the screening effects of UV-B effective for producing CPD levels and blue/UV-A effective for CPD photolyase activity.