Nucleic acid-binding properties of low-density lipoproteins: LDL as a natural gene vector
Nucleic acid-binding properties of low-density lipoproteins: LDL as a natural gene vector
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DOI：
10.1023/a:1020627212272
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发表时间：
1999-11-01
期刊：
影响因子：
--
通讯作者：
Moore, JP
中科院分区：
文献类型：
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作者：
Guevara, JG;Kang, DC;Moore, JP
apo B-100 作为转录因子的作用体现在其一级结构中存在与转录因子 ISGF3 gamma、STAT、IRF 和 SREBP 的 DNA 结合结构域相似的区域，以及 11 个 RNA 结合 KH 结构域的存在。 Apo B-100 序列还包含许多二分核定位序列 (NLS)。使用改良的凝胶位移测定来显示高度纯化的人 LDL 制剂与片段化基因组 DNA、质粒 DNA、合成寡核苷酸（ISRE、5'-GGGAAACC-GAAACTG 和 E/C、E-box 基序和 CCAAT、脂肪细胞特异性基因启动子位点）以及来自人肝脏的总 RNA 的结合。观察到 LDL 优先结合含有 hCMV IE2 启动子区域的质粒 DNA。在使用人肝脏总 RNA 的实验中，从 LDL 和 VLDL 条带中回收了五种不同基因的 RNA。使用人皮肤成纤维细胞进行基因转染实验来研究LDL的基因转移能力。如荧光所示，用 pEGFP-N1 质粒 DNA 和 LDL 转染的细胞在转染后约 3 小时表达功能性 GFP。我们的结果强烈支持 apo B-100 在控制基因表达和作为基因转移载体中的替代作用，无论是手提包还是功能片段。
The role of apo B-100 as a transcription factor is indicated by the presence of regions in its primary structure that are similar to the DNA-binding domains of the transcription factors ISGF3 gamma, STATs, IRFs, and SREBPs as well as by the presence of 11 RNA-binding KH domains. The Apo B-100 sequence also contains numerous bipartite nuclear localization sequences (NLS). A modified gel shift assay was used to show binding of highly purified preparations of human LDLs to fragmented genomic DNA, plasmid DNA, synthetic oligonucleotides (ISRE, 5'-GGGAAACC-GAAACTG and E/C, E-box motif and CCAAT, adipocyte-specific genes promoter site), and total RNA from human liver. LDL was observed to bind preferentially to plasmid DNA containing the hCMV IE2 promoter region. In experiments using human liver total RNA, RNA for five different genes was recovered from LDL and VLDL bands. Gene transfection experiments using human skin fibroblast cells were used to study the gene transfer capacity of LDL. Cells transfected with a pEGFP-N1 plasmid DNA and LDL expressed functional GFP, as indicated by fluorescence, at approximately 3 hrs after transfection. Our results strongly support an alternative role for apo B-100, in tote or perhaps as functional fragments, in the control of gene expression and as gene transfer vector.