Xiao L, Casero R A
Johns Hopkins Oncology Center Research Laboratories, Johns Hopkins University School of Medicine, Baltimore, MD 21231, USA.
Biochem J. 1996 Jan 15;313 ( Pt 2)(Pt 2):691-6. doi: 10.1042/bj3130691.
The expression of spermidine/spermine N1-acetyltransferase (SSAT), the rate-limiting enzyme in the catabolism of polyamines, is highly regulated by a number of factors including the natural polyamines and their analogues. The phenotype-specific cytotoxicity that occurs in response to a class of polyamine analogues, the diethylpolyamines, is associated with a phenotype-specific superinduction of SSAT in human non-small-cell lung carcinomas, whereas in non-responding cell types, including the small-cell lung carcinomas, the superinduction of SSAT does not occur. In this study, we have investigated the molecular basis of this phenotype-specific SSAT induction in human lung carcinoma cells in response to N1,N12-diethylspermine (BESpm). To facilitate the study of transcriptional regulation, we have cloned and characterized 11 kb of the human SSAT locus, including 3500 bp of the 5' promoter region. Nuclear run-on transcription studies suggest that the initial induction of SSAT results from an increase in the rate of gene transcription. Results from Northern blot analysis and ribonuclease protection assays indicate a differential expression of SSAT mRNA between the analogue-responsive H157 and non-responsive H82 cells. There is no detectable SSAT mRNA in H82 cells, even after a 24-h analogue treatment, whereas SSAT mRNA in H157 cells was detectable by Northern blot analysis and increased more than 100-fold following drug exposure. Furthermore, nuclear run-on transcription assays do not detect any active transcription of SSAT gene in either treated or untreated H82 cells. These results indicate that at least one component of the phenotype-specific induction of SSAT appears to be due to differences in transcriptional regulation of the gene. In addition, mapping of DNase I-hypersensitive sites of the SSAT gene suggest that the cell type-specific promoter/enhancer utilization may control the expression of the SSAT gene in differentially sensitive cell types in vivo.
多胺分解代谢中的限速酶——亚精胺/精胺N1 - 乙酰转移酶（SSAT）的表达受到多种因素的高度调控，这些因素包括天然多胺及其类似物。一类多胺类似物二乙多胺所引发的表型特异性细胞毒性，与人类非小细胞肺癌中SSAT的表型特异性超诱导相关，而在无反应的细胞类型中，包括小细胞肺癌，SSAT的超诱导则不会发生。在本研究中，我们调查了人类肺癌细胞中这种针对N1,N12 - 二乙亚精胺（BESpm）的表型特异性SSAT诱导的分子基础。为便于研究转录调控，我们克隆并鉴定了11 kb的人类SSAT基因座，包括5'启动子区域的3500 bp。细胞核连续转录研究表明，SSAT的初始诱导源于基因转录速率的增加。Northern印迹分析和核糖核酸酶保护试验的结果表明，在对类似物有反应的H157细胞和无反应的H82细胞之间，SSAT mRNA存在差异表达。即使经过24小时的类似物处理，H82细胞中也检测不到SSAT mRNA，而通过Northern印迹分析可检测到H157细胞中的SSAT mRNA，且在药物暴露后增加了100多倍。此外，细胞核连续转录试验在处理或未处理的H82细胞中均未检测到SSAT基因的任何活性转录。这些结果表明，SSAT表型特异性诱导的至少一个组成部分似乎是由于该基因转录调控的差异。此外，SSAT基因的DNase I超敏位点图谱表明，细胞类型特异性启动子/增强子的利用可能在体内差异敏感细胞类型中控制SSAT基因的表达。
