Mutagenic potential of benzo[a] pyrene-derived DNA adducts positioned in codon 273 of the human P53 gene
Mutagenic potential of benzo[a] pyrene-derived DNA adducts positioned in codon 273 of the human P53 gene
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DOI：
10.1021/bi0482194
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发表时间：
2004-12-21
期刊：
影响因子：
2.9
通讯作者：
Shibutani, S
中科院分区：
文献类型：
--
作者：
Dong, H;Bonala, RR;Shibutani, S
人类P53基因密码子273（（5’）CGT）是环境致癌物苯并[a]芘的突变热点。我们在人类P53基因密码子273的第二个位置加入了一个单(+)-或(-)-反式抗苯并[a]芘二醇环氧化物（BPDE） DNA加合物，并探索了这种病变在哺乳动物细胞中的致突变潜力。将(+)-或(-)-反式dg - n -2- bpde修饰的寡脱氧核糖核苷酸（5'GAGGTGCG(BIDE)TGTTTGT）整合到单链穿梭载体中，转染到猴肾细胞。然后用子代质粒转化大肠杆菌DH10B。利用寡脱氧核苷酸杂交和DNA序列分析对转化子进行分析，确定该加合碱基产生的突变频率和突变谱。我们确定与(+)-反式dg - n -2- bpde和(-)-反式dg - n -2- bpde内合相关的突变频率分别为26.5%和17.5%。这两种立体异构体产生的主要突变是G -> - T转换，还有一些G -> - A转换。当胞嘧啶5′to dG-N-2-BPDE被5-甲基胞嘧啶取代时，(+)-反式dg - n2 - bpde和(-)-反式dG-N-2-BPDE的突变频率分别降至11.1%和10.6%，突变特异性保持不变。因此，P53密码子273的突变“热点”可能反映了BPDE的序列特异性反应性和/或位于该位点的BPDE- dna加合物的低效修复。
Codon 273 ((5')CGT) of the human P53 gene is a mutational hot spot for the environmental carcinogen benzo[a]pyrene. We incorporated a single (+)- or (-)-trans-anti-benzo[a]pyrene diol epoxide (BPDE) DNA adduct at the second position of codon 273 of the human P53 gene and explored the mutagenic potential of this lesion in mammalian cells. Oligodeoxyribonucleotides (5'GAGGTGCG(BIDE)TGTTTGT) modified with (+)- or (-)-trans-dG-N-2-BPDE were incorporated into single-stranded shuttle vectors and transfected into simian kidney cells. Progeny plasmids were then used to transform Escherichia coli DH10B. Transformants were analyzed by oligodeoxynucleotide hybridization and DNA sequence analysis to establish the mutation frequency and spectrum produced by the adducted base. We determined the mutational frequencies associated with (+)-trans-dG-N-2-BPDE and (-)-trans-dG-N-2-BPDE adduction to be 26.5% and 17.5%, respectively. The predominant mutations generated by both stereoisomers were G --> T transversions, with some G --> A transitions. When the cytosine 5' to dG-N-2-BPDE was replaced by 5-methylcytosine, the mutational frequencies of (+)-trans-dG-N2-BPDE and (-)-trans-dG-N-2-BPDE were reduced to 11.1% and 10.6%, respectively, while the mutational specificity remained unchanged. Thus, the mutational "hot spot" at codon 273 in P53 may reflect either sequence-specific reactivity of BPDE and/or inefficient repair of BPDE-DNA adducts positioned at this site.