Regulation of the human T-cell receptor alpha gene enhancer: multiple ubiquitous and T-cell-specific nuclear proteins interact with four hypomethylated enhancer elements.
Regulation of the human T-cell receptor alpha gene enhancer: multiple ubiquitous and T-cell-specific nuclear proteins interact with four hypomethylated enhancer elements.
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人类 T 细胞受体 α 基因增强子的调节：多种普遍存在的 T 细胞特异性核蛋白与四种低甲基化增强子元件相互作用。
DOI：
10.1128/mcb.10.9.4720-4727.1990
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发表时间：
1990
影响因子：
5.3
通讯作者：
Leiden,JM
中科院分区：
文献类型：
--
作者：
Ho,IC;Leiden,JM
人T细胞受体（TCR）α基因的转录受T细胞特异性转录增强子调节，该增强子位于Ca基因片段的4.5个内切酶3'。以前的研究表明，这种增强子含有至少五个称为Tα1至Tα5的核蛋白结合位点。在本报告所述的研究中，我们确定了人TCR α增强子功能的分子要求。体外诱变和缺失分析表明，完整的增强子活性保留在含有Tal和Tα2核蛋白结合位点的116个碱基对片段中，并且这两个位点都是完整增强子功能所必需的。功能性增强子活性要求Tal和Tα2结合位点间隔大于15个且小于85个碱基对。然而，该间隔区的序列和DNA螺旋上两个结合位点的相对相位不影响增强子功能。缺失和突变分析表明，Tα3和Tα4核蛋白结合位点对TCR α增强子活性不是必需的或充分的。然而，含有这两个位点的片段能够补偿Tal和Tα2突变，否则会消除增强子活性。TCR α增强子结合蛋白的电泳迁移率变化分析显示，Tal、Ta3和Ta4结合蛋白在多种T细胞和非T细胞肿瘤细胞系中表达。相反，两个Ta2结合活性之一，仅在T细胞核提取物中检测到。TCR α增强子的活性似乎并非仅在DNA甲基化水平上受到调节，因为发现与成纤维细胞相比，增强子序列在B和T细胞中同样低甲基化。总之，这些结果表明，TCR α增强子活性受多种T细胞特异性和普遍存在的核蛋白与部分冗余顺式作用增强子元件（在淋巴系细胞中低甲基化）相互作用的调节。
Transcription of human T-cell receptor (TCR) α genes is regulated by a T-cell-specific transcriptional enhancer that is located 4.5 kilobases 3' of the Ca gene segment. Previous studies have demonstrated that this enhancer contains at least five nuclear protein-binding sites called Tα1 to Tα5. In the studies described in this report, we have determined the molecular requirements for human TCR α enhancer function. In vitro mutagenesis and deletion analyses demonstrated that full enhancer activity is retained in a 116-base-pair fragment containing the Tal and Tα2 nuclear protein-binding sites and that both of these sites are required for full enhancer function. Functional enhancer activity requires that the Tal and Tα2 binding sites be separated by more than 15 and fewer than 85 base pairs. However, the sequence of this spacer region and the relative phase of the two binding sites on the DNA helix do not affect enhancer function. Deletion and mutation analyses demonstrated that the Tα3 and Tα4 nuclear protein-binding sites are not necessary or sufficient for TCR α enhancer activity. However, a fragment containing these two sites was able to compensate for Tal and Tα2 mutations that otherwise abolished enhancer activity. Electrophoretic mobility shift analyses of the TCR α enhancer binding proteins revealed that the Tal, Ta3, and Ta4 binding proteins are expressed in a variety of T-cell and non-T-cell tumor cell lines. In contrast, one of the two Ta2 binding activities was detected only in T-cell nuclear extracts. The activity of the TCR α enhancer does not appear to be regulated solely at the level of DNA methylation in that the enhancer sequences were found to be identically hypomethylated in Β and Τ cells as compared with fibroblasts. Taken together, these results suggest that TCR α enhancer activity is regulated by the interaction of multiple T-cell-specific and ubiquitous nuclear proteins with partially redundant cis-acting enhancer elements that are hypomethylated in cells of the lymphoid lineage.