Parrish D D, Lambert M W
Department of Pathology, UMDNJ-New Jersey Medical School, Newark 07103.
Mutat Res. 1990 Mar;235(2):65-80. doi: 10.1016/0921-8777(90)90059-e.
The influence of nucleosome structure on the activity of 2 chromatin-associated DNA endonucleases, pIs 4.6 and 7.6, from normal human and xeroderma pigmentosum, complementation group A (XPA), lymphoblastoid cells was examined on DNA containing either psoralen monoadducts or cross-links. As substrate a reconstituted nucleosomal system was utilized consisting of a plasmid DNA and either core (H2A, H2B, H3, H4), or total (core plus H1) histones from normal or XPA cells. Both non-nucleosomal and nucleosomal DNA were treated with 8-methoxypsoralen (8-MOP) plus long-wavelength ultraviolet radiation (UVA), which produces monoadducts and DNA interstrand cross-links, and angelicin plus UVA, which produces monoadducts. Both normal endonucleases were over 2-fold more active on both types of psoralen-plus-UVA-damaged core nucleosomal DNA than on damaged non-nucleosomal DNA. Addition of histone H1 to the system reduced but did not abolish this increase. By contrast, neither XPA endonuclease showed any increase on psoralen-treated nucleosomal DNA, with or without histone H1. Mixing the normal with the XPA endonucleases led to complementation of the XPA defect. These results indicate that interaction of these endonucleases with chromatin is of critical importance and that it is at this level that a defect exists in XPA endonucleases.
研究了核小体结构对来自正常人及着色性干皮病A组（XPA）淋巴母细胞的两种染色质相关DNA内切酶（pI分别为4.6和7.6）活性的影响，这两种酶作用于含有补骨脂素单加合物或交联的DNA。作为底物，使用了一种重组核小体系统，该系统由质粒DNA以及来自正常或XPA细胞的核心组蛋白（H2A、H2B、H3、H4）或全部组蛋白（核心组蛋白加H1）组成。非核小体DNA和核小体DNA均用8-甲氧基补骨脂素（8-MOP）加长波紫外线（UVA）处理，后者可产生单加合物和DNA链间交联，以及用当归素加UVA处理，后者可产生单加合物。两种正常内切酶对两种类型的补骨脂素加UVA损伤的核心核小体DNA的活性均比对损伤的非核小体DNA的活性高2倍以上。向系统中添加组蛋白H1可降低但并未消除这种增加。相比之下，无论有无组蛋白H1，XPA内切酶对补骨脂素处理的核小体DNA均未表现出任何活性增加。将正常内切酶与XPA内切酶混合可导致XPA缺陷的互补。这些结果表明，这些内切酶与染色质的相互作用至关重要，并且正是在这个水平上，XPA内切酶存在缺陷。
