hSmad5 gene, a human hSmad family member:: its full length cDNA, genomic structure, promoter region and mutation analysis in human tumors
hSmad5 gene, a human hSmad family member:: its full length cDNA, genomic structure, promoter region and mutation analysis in human tumors
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DOI：
10.1038/sj.onc.1201614
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发表时间：
1998-02-19
期刊：
影响因子：
8
通讯作者：
Harris, CC
中科院分区：
文献类型：
--
作者：
Gemma, A;Hagiwara, K;Harris, CC
hSmad（mothers against decapentaplegic）相关蛋白是转化生长因子-β（1）（TGF-β（1））超家族信号转导途径中的重要信使。为了进一步鉴定该家族的成员，我们通过cDNA末端快速扩增（RACE）获得了人hSmad 5（hSmad 5）基因的全长cDNA，然后确定了该基因的基因组结构。有八个外显子和两个选择性转录本;较短的转录本缺少外显子2。我们从人基因组YAC克隆中鉴定了hSmad 5启动子区，通过获得cDNA 5'端上游延伸1235个碱基对的核苷酸序列。我们发现了一个CpG岛与启动子区域一致，我们证明了启动子活性在一个1232 bp的片段位于上游的转录起始位点。为了研究人类肿瘤中hSmad 5体细胞突变的频率，我们设计了基于内含子的引物，通过聚合酶链反应-单链构象多态性（PCR-SSCP）分析来检测编码区。在40个原发性胃肿瘤和51个来源于不同类型人类癌症的细胞系（包括20个对TGF-β的生长抑制作用具有抗性的细胞系）中均未发现纯合缺失或点突变（1）。这些结果表明，hSmad 5基因通常不会发生突变，其他遗传变异介导了人类癌症中TGF-β 1反应性的丧失。
hSmad (mothers against decapentaplegic)-related proteins are important messengers within the Transforming Growth Factor-beta(1) (TGF-beta(1)) superfamily signal transduction pathways. To further characterize a member of this family, we obtained a full length cDNA of the human hSmad5 (hSmad5) gene by rapid amplification of cDNA ends (RACE) and then determined the genomic structure of the gene. There are eight exons and two alternative transcripts; the shorter transcript lacks exon 2. We identified the hSmad5 promoter region from a human genomic YAC clone by obtaining the nucleotide sequence extending 1235 base pairs upstream of the 5' end of the cDNA. We found a CpG island consistent with a promoter region, and we demonstrated promoter activity in a 1232 bp fragment located upstream of the transcription initiation site. To investigate the frequency of somatic hSmad5 mutations in human cancers, we designed intron-based primers to examine coding regions by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis. Neither homozygous deletions or point mutations were found in 40 primary gastric tumors and 51 cell lines derived from diverse types of human cancer including 20 cell lines resistant to the growth inhibitory effects of TGF-beta(1). These results suggest that the hSmad5 gene is not commonly mutated and that other genetic alterations mediate the loss of TGF-beta(1) responsiveness in human cancers.