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肝癌抑制基因-1转运功能区的研究

A Study on Functional Region for Transportation in HCCS1
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摘要 肝癌抑制基因-1(hepatocellular carcinoma suppressor 1,HCCS1)是一种潜在的肝癌抑制基因,在细胞内发挥蛋白分拣运输的作用,其抑癌作用有可能与蛋白转运的功能有一定的关系。本研究旨在寻找HCCS1序列中与转运功能相关的序列区域。首先通过亚克隆技术,构建以pEGFP-C2为载体,对不同长度HCCS1-cDNA片段进行亚克隆,将构建的亚克隆转染到HeLa细胞,通过免疫荧光显微镜观察不同长度HCCS1蛋白在细胞内的分布,及其与6-磷酸甘露糖受体(M6PR)的共定位。经过PCR及亚克隆后,目的片段成功插入载体质粒,构建了以pEGFP-C2为载体的含有4个不同长度HCCS1片段的亚克隆。分别是:pEGFP-C2-N1836,pEGFP-C2-N1572,pEGFP-C2-1707和pEGFP-C2-1506,其中pEGFP-C2-N1836和pEGFP-C2-N1572编码的HCCS1蛋白,长度不同均呈颗粒状、极性分布于核周的胞质内,且与M6PR有共定位;而pEGFP-C2-1707片段编码的HCCS1蛋白虽然也呈颗粒状分布于核周,但极性分布消失,且与M6PR共定位消失;pEGFP-C2-1506与pEGFP-C2-1707结果相似。结果显示了HCCS1-cDNA的5’端的129 bp序列为与HCCS1的极性及M6PR共定位相关的区域。 Hepatocellular carcinoma suppressor gene-1（HCCS1） as a potential hepatocellular carcinoma supressor gene plays an important role in sorting of some cytoplasmic proteins.Its carcinoma suppressor function may be related to its protein import to some extent.So it is crucial to identify functional sequences in HCCS1 that are responsible for the transportation.The expression vectors containing different lengths of HCCS1 gene are constructed and transfected into HeLa cells mediated by liposomes.The localizations and co-localizations of different HCCS1 fragments are determined by fluorescence microscopy,respectively.Research indicates the polarity and co-localizations with M6PR both disappear when HCCS1 gene is cleaved 129 bp from the 5＇end by fluorescence microscopy.The results show that a 129 bp fragment in HCCS1 at the 5＇ end must be the region to its polarity and localization.
作者 周华 刘鸿 赵新泰 周秀梅
机构地区 浙江理工大学生命科学学院 浙江大学医学院 上海市肿瘤研究所
出处 《浙江理工大学学报（自然科学版）》 2011年第2期241-245,282,共6页 Journal of Zhejiang Sci-Tech University(Natural Sciences)
基金 国家自然科学基金(30623033)
关键词 肝癌抑制基因 功能序列 免疫荧光 转运 hepatocellular carcinoma supressor gene functional sequence immunofluorescence transportation
分类号 Q789 [生物学—分子生物学]
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参考文献12

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二级参考文献10

  • 1Zhao XT, Li JJ, He YH, et al. A novel growth suppressor gene on chromosome 17p13.3 with a high frequency of mutation in human hepatocellular carcinoma [ J ]. Cancer Res, 2001,61 ( 20 ) : 7383-7387.
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  • 3Conibear E,Stevens TH. Vps52p, Vps53p, and Vps54p form a novel multisubunit complex required for protein sorting at the yeast late Golgi [ J ]. Mol Biol Cell ,2000,11 ( 1 ) :305-323.
  • 4Conibear E, Stevens TH. Studying yeast vacuoles [ J ]. Methods Enzymol,2002 ,351:408-432.
  • 5Andag U, Neumann T, Schmitt HD. The coatomer-interacting protein Dsll p is required for Golgi-to-endoplasmic reticulum retrieval in yeast[ J]. Biol Chem,2001,276(42) :39150-39160.
  • 6Conibear E, Cleck JN, Stevens TH. Vps51p mediates the association of the GARP (Vps52/53/54) complex with the late Golgi t-SNARE Tlg1 p [ J ]. Mol. Biol. Cell,2003,14 (4) : 1610-1623.
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浙江理工大学学报（自然科学版）
2011年 第2期

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