Reed S H, McCready S, Boiteux S, Waters R
School of Biological Sciences, University College of Swansea, Swansea, United Kingdom.
Mol Gen Genet. 1996 Mar 7;250(4):515-22. doi: 10.1007/BF02174040.
In the accompanying paper we demonstrated that endonuclease III-sensitive sites in the MAT alpha and HML alpha loci of Saccharomyces cerevisiae are repaired by the Nucleotide Excision Repair (NER) pathway. In the current report we investigated the repair of endonuclease III sites, 6-4 photoproducts and cyclobutane pyrimidine dimers (CPDs) in a rad14-2 point mutant and in a rad14 deletion mutant. The RAD14 gene is the yeast homologue of the human gene that complements the defect in cells from xeroderma pigmentosum (XP) patients belonging to complementation group A. In the point mutant we observed normal repair of endonuclease III site (i.e. as wild type), but no removal of CPDs at the MAT alpha and HML alpha loci. Similar experiments were undertaken using the recently created rad14 deletion mutant. Here, neither endonuclease III sites nor CPDs were repaired in MATa or HMRa. Thus the point mutant appears to produce a gene product that permits the repair of endonuclease III sites, but prevents the repair of CPDs. Previously it was found that in the genome overall, repair of 6-4 photoproducts was less impaired that repair of CPDs in the point mutant. The deletion mutant repairs neither CPDs nor 6-4 photoproducts in the genome overall. This finding is consistent with the RAD14 protein being involved in lesion recognition in yeast. A logical interpretation is that the rad14-2 point mutant produces a modified protein that enables the cell to repair endonuclease III sites and 6-4 photoproducts much more efficiently than CPDs. This modified protein may aid studies designed to elucidate the role of the RAD14 protein in lesion recognition.
在随附论文中，我们证明了酿酒酵母MATα和HMLα基因座中的核酸内切酶III敏感位点可通过核苷酸切除修复（NER）途径进行修复。在本报告中，我们研究了rad14 - 2点突变体和rad14缺失突变体中核酸内切酶III位点、6 - 4光产物和环丁烷嘧啶二聚体（CPD）的修复情况。RAD14基因是人类基因的酵母同源物，该人类基因可弥补属于A互补组的着色性干皮病（XP）患者细胞中的缺陷。在点突变体中，我们观察到核酸内切酶III位点的修复正常（即与野生型一样），但在MATα和HMLα基因座处未检测到CPD的去除。使用最近创建的rad14缺失突变体进行了类似实验。在此，MATa或HMRa中核酸内切酶III位点和CPD均未得到修复。因此，点突变体似乎产生了一种基因产物，该产物允许核酸内切酶III位点的修复，但阻止了CPD的修复。此前发现，在整个基因组中，点突变体中6 - 4光产物的修复受损程度低于CPD的修复。缺失突变体在整个基因组中既不修复CPD也不修复6 - 4光产物。这一发现与RAD14蛋白参与酵母中的损伤识别一致。一个合理的解释是，rad14 - 2点突变体产生了一种修饰蛋白，使细胞能够比修复CPD更有效地修复核酸内切酶III位点和6 - 4光产物。这种修饰蛋白可能有助于旨在阐明RAD14蛋白在损伤识别中作用的研究。
