NONRANDOM BINDING OF THE CARCINOGEN N-HYDROXY-2-ACETYLAMINOFLUORENE TO REPETITIVE SEQUENCES OF RAT-LIVER DNA INVIVO
NONRANDOM BINDING OF THE CARCINOGEN N-HYDROXY-2-ACETYLAMINOFLUORENE TO REPETITIVE SEQUENCES OF RAT-LIVER DNA INVIVO
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DOI：
10.1073/pnas.81.22.6943
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发表时间：
1984-01-01
期刊：
影响因子：
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通讯作者：
GUPTA, RC
中科院分区：
文献类型：
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作者：
GUPTA, RC
在大鼠肝脏中，研究了单剂量致癌物N-羟基-2-乙酰氨基芴后，单个加合物在重复DNA序列中的分布。通过用HindIII限制性内切酶（EC 3.1.23.21）消化肝DNA和凝胶电泳分离重复片段[82、125、179、225和370个碱基对（bp）]。作为32 P postlabeling测定，没有观察到重复序列和总DNA的DNA结合代谢之间的定性差异，但优先结合这些序列发生。处理1天后，N-羟基-2-乙酰氨基芴诱导的加合物的量在179、225和370 bp重复序列中分别比总DNA高13.8、2.0和3.0倍，而82和125 bp重复序列没有显示出差异。各个加合物的相对分布在各种序列之间变化。9天后，与总DNA相比，所有5个序列显示出1.3-1.7倍的结合。相反，当DNA在体外与活性代谢物N-乙酰氧基-2-乙酰氨基芴反应时，观察到随机结合。这些结果表明，在重复DNA序列中的加合物的富集和差异切除可能是DNA的核组织的功能。32 P测定的这种应用构成了以比迄今为止可能的更灵敏和精确的方式研究染色质结构组分（包括转录和非转录的多拷贝基因）中体内DNA损伤和切除修复的手段。
The distribution of individual adducts in repetitive DNA sequences of rat liver was examined in vivo after a single dose of the carcinogen N-hydroxy-2-acetylaminofluorene. Repetitive fragments [82, 125, 179, 225 and 370 base pairs (bp)] were isolated by digestion of hepatic DNA with HindIII restriction endonuclease (EC 3.1.23.21) and gel electrophoresis. As assayed by 32P postlabeling, no qualitative differences were observed between the DNA-bound metabolities in repetitive sequences and total DNA, but preferential binding to these sequences occurred. After 1 day of treatment, the amounts of N-hydroxy-2-acetylaminofluorene-induced adducts were 13.8, 2.0 and 3.0 times higher in 179, 225 and 370 bp repeats, respectively, than in total DNA, while 82 and 125 bp repeats showed no differences. The relative distribution of individual adducts varied among the various sequences. After 9 days, all 5 sequences showed 1.3-1.7 times higher binding as compared to total DNA. In contrast, a random binding was observed when DNA reacted in vitro with an active metabolite, N-acetoxy-2-acetylaminofluorene. These results suggest that the enrichment and differential excision of adducts in the repetitive DNA sequences may be a function of the nuclear organization of DNA. This application of the 32P assay constitutes a means to study the DNA damage and excision repair in vivo in chromatin structural components, including transcribed and nontranscribed multiple-copy genes in a much more sensitive and precise way than has been hitherto possible.