Structure and transcriptional regulation of the Nat2 gene encoding for the drug-metabolizing enzyme arylamine N-acetyltransferase type 2 in mice
Structure and transcriptional regulation of the Nat2 gene encoding for the drug-metabolizing enzyme arylamine N-acetyltransferase type 2 in mice
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DOI：
10.1042/bj20030812
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发表时间：
2003-11-01
影响因子：
4.1
通讯作者：
Sim, E
中科院分区：
文献类型：
--
作者：
Boukouvala, S;Price, N;Sim, E
芳香胺N-乙酰基转移酶（NAT）是一种多态性酶，在药物和致癌物的代谢中起重要作用。小鼠有三种NAT同工酶，其中NAT 2被假定参与内源性代谢以及外源性代谢。为了了解小鼠Nat 2基因的表达，我们分析了其结构和转录调控。我们已经准确地映射了转录起始位点6.5 kb的基因编码区上游，邻近最近描述的非编码外显子。在胚胎和成体肝、脾、颌下腺、肾、脑、胸腺、肺和胎盘中，转录被证明是从该区域开始的，但在心脏中不是。通过PCR对cDNA进行数据库搜索和分析，表明Nat 2的单个6.2 kb内含子的选择性剪接，并确定了该基因编码区下游0.44 kb处的聚腺苷酸化信号的位置。检查侧翼的13 kb序列的编码和非编码外显子的Nat 2揭示了一个单一的启动子，位于转录起始位点附近，并表示可能的海港控制元件的地区。Nat 2启动子由一个非典型的TATA盒和一个Sp1 [SV 40（猿猴病毒40）蛋白1]盒组成，与许多管家基因启动子中发现的相同。Nat 2启动子的活性被这两个元素中的任一个的缺失或突变严重降低，而Sp1盒的区域结合细胞蛋白并抵抗DNase I消化。最后，启动子区域结合细胞蛋白的能力通过与具有Sp1共有序列的寡核苷酸竞争而降低。
Arylamine N-acetyltransferases (NATs) are polymorphic enzymes, well-known for their role in the metabolism of drugs and carcinogens. Mice have three NAT isoenzymes, of which NAT2 is postulated to be involved in endogenous, as well as xenobiotic, metabolism. To understand expression of the murine Nat2 gene, we have analysed its structure and transcriptional regulation. We have accurately mapped the transcription initiation site 6.5 kb upstream of the coding region of the gene, adjacent to a recently described non-coding exon. Transcription was demonstrated to start from this region in embryonic and adult liver, spleen, submaxillary gland, kidney, brain, thymus, lung and placenta, but not in the heart. Database searches and analyses of cDNA by PCR suggested alternative splicing of the single 6.2 kb intron of Nat2, and determined the position of the polyadenylation signal at 0.44 kb downstream of the coding region of the gene. Examination of the 13 kb sequence flanking the coding and non-coding exons of Nat2 revealed a single promoter, located close to the transcription-initiation site, and indicated regions likely to harbour control elements. The Nat2 promoter consists of an atypical TATA box and a Sp1 [SV40 (simian virus 40) protein 1] box identical with that found in many housekeeping gene promoters. Activity of the Nat2 promoter was severely reduced by deletion or mutation of either of these two elements, whereas the region of the Sp1 box bound cellular protein and resisted DNase I digestion. Finally, the ability of the promoter region to bind cellular protein was reduced by competition with oligonucleotides bearing the Sp1 consensus sequence.