【作者】 周津；

【导师】 吴旻；

【作者基本信息】 中国协和医科大学 ， 细胞生物学， 2000， 博士

【摘要】 为了解食管癌发生的分子遗传学机理，分离食管癌中差异表达的基因，本研究应用抑制消减杂交并与PCR合成双链cDNA、反Northern筛选SSH克隆相结合，自微克级总RNA中分离出多个食管癌中差异表达的基因，其中表达明显下调的Lipocortin I、Cystatin A、Cystatin B、Cytokeratin 13等可能与食管癌的去分化、恶性增殖转化有关。应用RACE等方法，获得了一个全长866bp的新基因，命名为NMES1(Normal mucosa of esophagus specific 1)，编码一个含83个氨基酸的蛋白质。Northern及RT-PCR分析显示该基因在食管癌中表达明显下调，应用RH定位的方法，NMES1定位于15q21.1。蛋白质序列同源分析显示可能与NADH：泛醌氧化还原酶MLRQ亚基(CI-MLRQ)功能相关，但不属于氧化还原呼吸链的组分，以NMES1转染人食管癌细胞系9706,可导致细胞生长减缓，显示该基因可能在食管、胃、小肠、结肠等消化道粘膜具有特殊的功能，且与食管癌的发生发展有关。 为全面了解癌与癌旁组织以及从正常食管粘膜、癌变过程中的各个阶段到食管癌发生基因表达谱的改变，应用近年来发展的微阵列技术，分析和比较了588个肿瘤相关基因在食管癌和对应癌旁组织，以及在正常成人食管粘膜、基底细胞增生Ⅱ级、重度不典型增生、原位癌、早期侵润癌组织中的表达状况，结果发现从正常食管粘膜发展成食管癌的各个阶段都有上百个基因表达发生变化，而且基因表达谱在癌变过程中随着细胞表型的演变而变化。这些基因及其在一定时期、一定组织中的表达变化组成了癌与癌旁组织，及各更多还原

【Abstract】 In an effort to isolate genes with different expression level in esophageal cancer, we performed suppression subtractive hybridization in combination with PCR-based cDNA synthesis and reverse Northern on the cancer tissues and matched adjacent normal mucosa using 5 microgram of total RNA as starting material. Among the differentially expressed genes identified, Lipocortin I、 Cystatin A、 Cystatin B、 Cytokeratin 13 are down regulated substantially. Also a cDNA ful-length of 866bp corresponding to a novel gene has been obtained by RACE, expression of which was observed in 11/12 adjacent normal esophageal mucosa, while the matched cancer tissues showed a drastic reduction or a complete lack of its expression. The corresponding gene is designated NMES1 (normal mucosa of esophagus specific 1). Sequence analysis of its full-length cDNA revealed a novel gene with a predicted polypeptide of 9 kD and sharing a 31 ~34% similarity with the NADH.ubiquinone oxidoreductase MLRQ subunit in mouse, bovine and human. Northern blot showed that NMES1 distributed mainly in alimentary tract, such as esophagus, stomach, small intestine, and colon. Expression in other tissues tested was only observed in placenta and lung, which is quite different from that of the members of mitochondrial respiratory chain which are always present in all tissues, with higher expression level in heart, skeletal更多还原