6q25区域内一个新基因MTLC的克隆及特性分析

Cloning and characterization of MTLC, a novel gene in 6q25

目的 克隆 6q2 5区域内喉癌相关新基因。方法 以表达序列标签为电子探针 ,在人类基因组数据库中进行电子杂交 ,钓出相应基因组DNA克隆后进行基因预测。根据获得的基因序列设计引物 ,逆转录 聚合酶链反应扩增新基因cDNA。结果 克隆了 1个 6q2 5区域内的新基因。该基因全长约 2 1kb ,含两个外显子 ,其cDNA序列长 10 0 6bp ,编码蛋白包括 2 3 5个氨基酸。其 5′侧翼序列存在癌蛋白c Myc的结合位点 ,将其命名为MTLC (c Myctargetfromlaryngealcancercells)基因。同源性分析表明该基因编码蛋白与小鼠MT MC1蛋白同源性达 78%。MTLC蛋白一级结构含有一个核定位信号结构域 ,亚细胞定位实验证实该基因主要在细胞核表达 ,Northern印迹分析表明MTLC基因在心肌、肝、肾、脑等多个组织有表达。结论 成功克隆了 1个新基因MTLC ,该基因可能作为c Myc的靶基因以转录因子形式参与维持细胞正常生理功能。

Objective: To identify and characterize laryngeal cancer related novel genes located on chromosome 6q25. Methods: Electric hybridization was performed in human genome database using EST (expression sequence tag) as probe. Novel genes were deduced by software from positive DNA clones and their cDNAs were amplified by RT-PCR using primers designed according to the sequence of the putative genes. Results: A novel gene was cloned successfully. The full length of this gene was about 21 kb. It contained two exons and produced a 1006 bp transcript coding a protein with 235 amino acid residues. It's 5′ flanking sequence contained two binding sites of oncoprotein c-Myc, thus it was named MTLC (c-Myc target from laryngeal cancer cells). Homologous assay showed that MTLC exhibited little overall homology to known human proteins but it exhibited good overall homology to mouse MT-MC1 protein with an identity of 78%. The primary structure of MTLC protein contained a nuclear location signal motif, but it did not have other conserved domains. The results of subcellular location experiment showed that MTLC expressed in nuclei of human hepatocellular carcinoma cell line Be17402 cells, while a wide distribution of MTLC in various tissues was demonstrated by Northern blotting. Conclusion: MTLC may play an important role as a target gene of c-Myc and as a transcription factor in keeping the normal physiological process of cells.