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目的 以往研究表明肝癌中染色体 1 7p1 3 .3区有高频率的杂合性缺失。其最小杂合性缺失范围已被确定在D1 7S643至D1 7S1 574位点间 ,而且其中的D1 7S92 6位点有最高的杂合性缺失率。含有该位点的基因组克隆P579已被测序分析 ,在P579范围共有 1 3个新基因。这里报告其中的一个新基因 (命名为肝癌抑癌基因 1 ,HCCS1 )的克隆和特性研究结果。方法 利用直接杂交筛选方法获得基因组克隆P579中的基因克隆。根据HCCS1基因克隆的cDNA序列与基因组序列进行比较确定基因的外显子与内含子。应用RT PCR扩增组织中的HCCS1基因 ,序列测定检查突变。应用免疫组化检测HCCS1在组织中的表达。应用克隆形成试验和裸鼠成瘤试验检测HCCS1的生物学功能。结果 HCCS1有 1 8个外显子 ,cDNA全长约2 .0kb ,蛋白产物定位于线粒体。HCCS1在肝癌组织中有高频率的突变 ,免疫组化检测表明HCCS1在癌旁组织的表达明显高于癌组织。HCCS1转染肝癌细胞明显抑制其克隆的形成及在裸鼠体内的成瘤。结论 上述发现表明HCCS1具有肝癌抑癌基因的作用
Purpose Previous studies have shown that there is a high frequency of loss of heterozygosity in chromosome 17pl3.3 in HCC. The minimum range of loss of heterozygosity has been identified between the D1 7S643 to D1 7S1 574 sites, and the D1 7S92 6 site has the highest loss of heterozygosity. The genomic clone P579 containing this site has been sequenced and analyzed, with a total of 13 new genes in the P579 range. Here we report the cloning and characterization of one of the new genes, named HCCS1. Methods The gene clone in genomic clone P579 was obtained by direct hybridization screening. The exon and intron of the gene were determined by comparing the cDNA sequence of the HCCS1 gene with the genomic sequence. The HCCS1 gene was amplified by RT PCR and sequenced for mutations. The expression of HCCS1 in tissues was detected by immunohistochemistry. The biological function of HCCS1 was detected by clonogenic assay and nude mouse tumorigenicity assay. Results There were 18 exons in HCCS1 and the cDNA was about 2.0 kb in length. The protein product was located in mitochondria. HCCS1 has a high frequency of mutations in HCC tissues. Immunohistochemistry showed that the expression of HCCS1 in adjacent tissues was significantly higher than that in cancer tissues. HCCS1 transfected hepatoma cells significantly inhibited the formation of its clone and tumor formation in nude mice. Conclusion The above findings indicate that HCCS1 has the function of tumor suppressor gene of liver cancer