Structure-function correlations of the insulin-linked polymorphic region
Structure-function correlations of the insulin-linked polymorphic region
复制标题
DOI：
10.1006/jmbi.1996.0659
复制
发表时间：
1996-12-06
影响因子：
5.6
通讯作者：
Gupta, G
中科院分区：
文献类型：
--
作者：
Catasti, P;Chen, X;Gupta, G
胰岛素连锁多态性区（insulin-linked polymorphism region，ILPR）的胰岛素小卫星位于人胰岛素基因上游363个碱基对处，是一个14个碱基对长的串联重复序列：5 '-ACAGGGGTGTGGGG-3' 3 '-TGTCCCCACACCCC-5'。胰岛素依赖型糖尿病（IDDM）的基因座已被定位到ILPR。ILPR基因的长度多态性与胰岛素基因的转录活性和胰岛素依赖型糖尿病的易感性有关。在这里，我们试图破译长度多态性和转录调控的ILPR结构的作用。我们通过凝胶电泳，圆二色性（CD）和一维和二维核磁共振光谱（1D/2D NMR），富含G链的ILPR采用分子内折叠发夹G-四重结构。对d（G（4）TGTG（4））和d（G（4）TGTG（4）ACAG（4）TGTG（4））的1D/2D NMR数据的详细分析使我们能够定义链折叠的性质、茎中G-四联体的堆积相互作用以及环中碱基的相互作用。d（G（4）TGTG（4）ACAG（4）TGTG（4））恰好是能够形成分子内发夹状G四联体结构的富G链的最小单元。对于长ILPR序列，几个这样的发夹G-四联体结构可以在空间中连接。事实上，通过体外复制测定，我们显示了重复长度为6的ILPR的富含G的链存在这样的多个发夹G-四联体结构。这一观察结果表明，多个发夹G-四联体的形成可以解释复制过程中的滑动和观察到的长度多态性。从我们的高分辨率结构中，我们能够确定一组对发夹G-四重峰的结构和稳定性至关重要的相互作用。ILPR中破坏这些相互作用的单突变或双突变也会降低胰岛素基因的转录活性。因此，ILPR的发夹G-四联体结构与人胰岛素基因的转录活性具有直接相关性。(C)1996年学术出版社
The insulin minisatellite of the insulin-linked polymorphic region (ILPR), a 14 base-pairs long tandem repeat of: 5'-ACAGGGGTGTGGGG-3' 3'-TGTCCCCACACCCC-5', is located 363 base-pairs upstream of the human insulin gene. A locus for insulin-dependent diabetes mellitus (IDDM) has been mapped to the ILPR. It has been shown that the ILPR is polymorphic in length and this length polymorphism is also related to the transcriptional activity of the insulin gene and the susceptibility to IDDM. Here, we attempt to decipher the role of the ILPR structure in length polymorphism and transcriptional regulation. We show by gel electrophoresis, circular dichroism (CD) and one and two-dimensional nuclear magnetic resonance spectroscopy (1D/2D NMR) that the G-rich strand of the ILPR adopts an intramolecularly folded hairpin G-quartet structure. A detailed analysis of 1D/2D NMR data of d(G(4)TGTG(4)) and d(G(4)TGTG(4)ACAG(4)TGTG(4)) enables us to define the nature of chainfolding, the stacking interaction of the G-tetrads in the stem, and the interactions of the bases in the loops. d(G(4)TGTG(4)ACAG(4)TGTG(4)) happens to be the smallest unit of the G-rich strand that can form the intramolecular hairpin G-quartet structure. For long ILPR sequences, several such hairpin G-quartet structures can be linked in space. Indeed, by an in vitro replication assay, we show the presence of such multiple hairpin G-quartet structures for the G-rich strand of the ILPR of repeat length 6. This observation suggests that the formation of multiple hairpin G-quartets may explain slippage during replication and the observed length polymorphism. From our high resolution structure, we are able to identify a set of interactions that are critical for the structure and stability of the hairpin G-quartet. Single or double mutations in the ILPR that destabilize these interactions also lower the transcriptional activity of the insulin gene. Therefore, the hairpin G-quartet structure of the ILPR has a direct correlation with the transcriptional activity of the human insulin gene. (C) 1996 Academic Press Limited