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首页> 外文期刊>Molecular and Cellular Biology >Variegated Expression from the Murine Band 3 (AE1) Promoter in Transgenic Mice Is Associated with mRNA Transcript Initiation at Upstream Start Sites and Can Be Suppressed by the Addition of the Chicken β-Globin 5′ HS4 Insulator Element
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Variegated Expression from the Murine Band 3 (AE1) Promoter in Transgenic Mice Is Associated with mRNA Transcript Initiation at Upstream Start Sites and Can Be Suppressed by the Addition of the Chicken β-Globin 5′ HS4 Insulator Element

机译：小鼠带3（AE1）启动子在转基因小鼠中的杂色表达与上游起始位点的mRNA转录起始相关，并且可以通过添加鸡β-球蛋白5'HS4绝缘子元件来抑制

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The anion exchanger protein 1 (AE1; band 3) is an abundant erythrocyte transmembrane protein that regulates chloride-bicarbonate exchange and provides an attachment site for the erythrocyte membrane skeleton on the cytoplasmic domain. We analyzed the function of the erythroid AE1 gene promoter by using run-on transcription, RNase protection, transient transfection, and transgenic mouse assays. AE1 mRNA was transcribed at a higher level and maintained at a higher steady-state level than either ankyrin or β-spectrin in mouse fetal liver cells. When linked to a human γ-globin gene, two different AE1 promoters directed erythroid-specific expression of γ-globin mRNA in 18 of 18 lines of transgenic mice. However, variegated expression of γ-globin was observed in 14 of 18 lines. While there was a significant correlation between transgene copy number and the amount of γ-globin mRNA in all 18 lines, the transgene mRNAs initiated upstream of the start site of the endogenous AE1 mRNA. Addition of the insulator element from 5′HS4 of the chicken β-globin cluster to the AE1/γ-globin transgene allowed position-independent, copy-number-dependent expression at levels similar to the AE1 transcription rate in six of six lines of transgenic mice. The mRNA from the insulated AE1/γ-globin transgene mapped to the start site of the endogenous AE1 mRNA, and γ-globin protein was expressed in 100% of erythrocytes in all lines. We conclude that the chicken β-globin 5′HS4 element is necessary for full function of the AE1 promoter and that position effect variegation is associated with RNA transcription from the upstream start sites.
机译：阴离子交换蛋白1（AE1;带3）是一种丰富的红细胞跨膜蛋白，可调节氯离子-碳酸氢根的交换，并为细胞质结构域上的红细胞膜骨架提供附着位点。我们通过使用连续转录，RNase保护，瞬时转染和转基因小鼠实验分析了类红细胞AE1基因启动子的功能。在小鼠胎儿肝细胞中，AE1 mRNA的转录水平高于锚蛋白或β-血影蛋白，并保持较高的稳态水平。当与人类γ-球蛋白基因连接时，两个不同的AE1启动子在18条转基因小鼠中的18条中指导γ-球蛋白mRNA的类红血球特异性表达。然而，在18个系中的14个中观察到γ-珠蛋白的杂色表达。在所有18个品系中，转基因拷贝数与γ-珠蛋白mRNA量之间存在显着相关性，而转基因mRNA在内源性AE1 mRNA起始位点上游开始。将鸡β-球蛋白簇的5'HS4的绝缘子元件添加到AE1 /γ-球蛋白转基因中，可以实现位置独立，拷贝数依赖的表达，其表达水平类似于六种转基因中的六种中的AE1转录速率老鼠。来自绝缘的AE1 /γ-球蛋白转基因的mRNA定位于内源性AE1 mRNA的起始位点，并且γ-球蛋白在所有系中的100％红细胞中表达。我们得出结论，鸡β-珠蛋白5'HS4元件对于AE1启动子的完整功能是必需的，并且位置效应的变异与上游上游位点的RNA转录有关。

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