Joe C O, Sylvia V L, Norman J O, Busbee D L
Mutat Res. 1987 Sep;184(2):129-37. doi: 10.1016/0167-8817(87)90069-1.
Normal human fibroblasts treated with r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) yielded DNA polymerase alpha with elevated levels of activity, incorporated [3H]thymidine as a function of unscheduled DNA synthesis, and exhibited restoration of normal DNA-strand length as a function of unscheduled DNA synthesis. Lipoprotein-deficient fibroblasts treated with BPDE did not show elevated levels of DNA polymerase alpha activity, exhibited minimal [3H]thymidine incorporation, and had fragmented DNA after 24 h of repair in the absence of lipoprotein or phosphatidylinositol supplementation. When DNA polymerase beta activity was inhibited, cells with normal lipoprotein uptake exhibited [3H]thymidine incorporation into BPDE-damaged DNA but did not show an increase in DNA-strand length. DNA polymerase alpha activity and [3H]thymidine incorporation in lipoprotein-deficient fibroblasts increased to normal levels when the cells were permeabilized and low-density lipoproteins or phosphatidylinositol were introduced into the cells. DNA polymerase alpha isolated from normal human fibroblasts, but not from lipoprotein-deficient fibroblasts, showed increased specific activity after the cells were treated with BPDE. When BPDE-treated lipoprotein-deficient fibroblasts were permeabilized and 32P-ATP was introduced into the cells along with lipoproteins, 32P-labeled DNA polymerase alpha with significantly increased specific activity was isolated from the cells. These data suggest that treatment of human fibroblasts with BPDE initiates unscheduled DNA synthesis, as a function of DNA excision repair, which is correlated with increased activity of DNA polymerase alpha, and that increased DNA polymerase alpha activity may be correlated with phosphorylation of the enzyme in a reaction that is stimulated by low-density lipoprotein or by the lipoprotein component, phosphatidylinositol.
用r-7,t-8-二羟基-t-9,10-环氧-7,8,9,10-四氢苯并[a]芘（BPDE）处理正常人类成纤维细胞后，产生了活性水平升高的DNA聚合酶α，其作为非预定DNA合成的函数掺入了[3H]胸苷，并随着非预定DNA合成表现出正常DNA链长度的恢复。用BPDE处理的脂蛋白缺陷型成纤维细胞未显示出DNA聚合酶α活性水平升高，表现出最小的[3H]胸苷掺入，并且在缺乏脂蛋白或磷脂酰肌醇补充的情况下修复24小时后DNA发生片段化。当DNA聚合酶β活性受到抑制时，具有正常脂蛋白摄取的细胞将[3H]胸苷掺入BPDE损伤的DNA中，但未显示出DNA链长度增加。当细胞被通透化并将低密度脂蛋白或磷脂酰肌醇引入细胞时，脂蛋白缺陷型成纤维细胞中的DNA聚合酶α活性和[3H]胸苷掺入增加到正常水平。从正常人类成纤维细胞而非脂蛋白缺陷型成纤维细胞中分离出的DNA聚合酶α，在用BPDE处理细胞后显示出比活性增加。当用BPDE处理的脂蛋白缺陷型成纤维细胞被通透化并将32P-ATP与脂蛋白一起引入细胞时，从细胞中分离出具有显著增加的比活性的32P标记的DNA聚合酶α。这些数据表明，用BPDE处理人类成纤维细胞可启动非预定DNA合成，这是DNA切除修复的函数，与DNA聚合酶α活性增加相关，并且DNA聚合酶α活性增加可能与该酶在由低密度脂蛋白或脂蛋白成分磷脂酰肌醇刺激的反应中的磷酸化相关。
