Teng B, Davidson N O
Department of Medicine, University of Chicago, Illinois 60637.
J Biol Chem. 1992 Oct 15;267(29):21265-72.
Mammalian intestinal apolipoprotein B (apoB) messenger RNA (mRNA) undergoes posttranscriptional editing, changing codon 2153 from CAA in apoB100 mRNA to an in-frame translational stop codon (UAA) in apoB48 mRNA. By contrast, chicken intestinal apoB cDNA contains a CAA codon at the corresponding site and apoB mRNA from chicken enterocytes, kidney, and liver is unedited. The cDNA sequence of chicken apoB spanning the edited base is divergent from mammalian apoB cDNA sequence, with 70% homology over the conserved 29-nucleotide sequence (6662-6690) flanking codon 2153. Efficient in vitro editing of both human and rat, but not chicken, synthetic apoB RNA was achieved using rat enterocyte S-100 extracts. By contrast, chicken enterocyte S-100 extracts failed to edit chicken, rat, or human synthetic apoB RNA. Mixing experiments, however, revealed that chicken enterocyte S-100 extracts enhance the in vitro editing activity of rat, pig, and human enterocyte S-100 extracts upon homologous RNAs. The editing enhancement activity of chicken enterocyte S-100 extracts is tissue-specific, heat-sensitive, substrate-saturable, and sensitive to proteinase K, but resistant to micrococcal nuclease. The activity was partially purified by Q-Sepharose chromatography and has an average molecular mass of 49 kDa when analyzed by gel filtration chromatography. We conclude that the evolutionary adaptation of intestinal apoB mRNA editing requires both a requisite RNA motif and tissue-specific factors which mediate the site-specific modification.
哺乳动物肠道载脂蛋白B(apoB)信使核糖核酸(mRNA)会经历转录后编辑,将apoB100 mRNA中的密码子2153从CAA变为apoB48 mRNA中的框内翻译终止密码子(UAA)。相比之下,鸡肠道apoB互补脱氧核糖核酸(cDNA)在相应位点含有CAA密码子,来自鸡肠细胞、肾脏和肝脏的apoB mRNA未被编辑。跨越编辑碱基的鸡apoB cDNA序列与哺乳动物apoB cDNA序列不同,在密码子2153侧翼保守的29个核苷酸序列(6662 - 6690)上有70%的同源性。使用大鼠肠细胞S - 100提取物实现了对人和大鼠而非鸡的合成apoB RNA的高效体外编辑。相比之下,鸡肠细胞S - 100提取物未能编辑鸡、大鼠或人的合成apoB RNA。然而,混合实验表明,鸡肠细胞S - 100提取物可增强大鼠、猪和人肠细胞S - 100提取物对同源RNA的体外编辑活性。鸡肠细胞S - 100提取物的编辑增强活性具有组织特异性、热敏感性、底物饱和性且对蛋白酶K敏感,但对微球菌核酸酶有抗性。该活性通过Q - 琼脂糖凝胶柱层析部分纯化,经凝胶过滤层析分析,其平均分子量为49 kDa。我们得出结论,肠道apoB mRNA编辑的进化适应既需要必需的RNA基序,也需要介导位点特异性修饰的组织特异性因子。