Requirement of dimerization for RNA editing activity of adenosine deaminases acting on RNA
Requirement of dimerization for RNA editing activity of adenosine deaminases acting on RNA
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DOI：
10.1074/jbc.m213127200
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发表时间：
2003-05-09
影响因子：
4.8
通讯作者：
Nishikura, K
中科院分区：
文献类型：
--
作者：
Cho, DSC;Yang, WD;Nishikura, K
作用于RNA的腺苷脱氨酶（阿达尔）将双链RNA中的腺苷残基转化为肌苷。已鉴定出三种脊椎动物阿达尔基因家族成员，ADAR 1、ADAR 2和ADAR 3。所有三个阿达尔基因家族成员的催化结构域与大肠杆菌胞苷脱氨酶和APOBEC-1的催化结构域非常相似。同二聚化对于这些胞苷脱氨酶的酶活性是必不可少的。在这项研究中，我们通过连续亲和层析和尺寸排阻柱层析研究了差异表位标记的阿达尔单体之间复合物的形成。ADAR 1和ADAR 2形成稳定的酶活性同源二聚体复合物，而ADAR 3保持为单体的酶失活形式。未检测到不同阿达尔基因家族成员之间形成异二聚体复合物。HeLa和小鼠脑核提取物的分析表明，内源性ADAR 1和ADAR 2两者形成同源二聚体复合物。有趣的是，内源性ADAR 3似乎也形成同源二聚体复合物，表明ADAR 3二聚化存在脑特异性机制。同二聚体的形成可能是阿达尔发挥活性脱氨酶作用所必需的。由一个野生型和一个突变体单体组成的二聚体复合物的分析表明，在位点选择性RNA编辑过程中，两个亚基之间存在功能性相互作用。
Adenosine deaminases acting on RNA (ADAR) convert adenosine residues into inosines in double-stranded RNA. Three vertebrate ADAR gene family members, ADAR1, ADAR2, and ADAR3, have been identified. The catalytic domain of all three ADAR gene family members is very similar to that of Escherichia coli cytidine deaminase and APOBEC-1. Homodimerization is essential for the enzyme activity of those cytidine deaminases. In this study, we investigated the formation of complexes between differentially epitope-tagged ADAR monomers by sequential affinity chromatography and size exclusion column chromatography. Both ADAR1 and ADAR2 form a stable enzymatically active homodimer complex, whereas ADAR3 remains as a monomeric, enzymatically inactive form. No heterodimer complex formation among different ADAR gene family members was detected. Analysis of HeLa and mouse brain nuclear extracts suggested that endogenous ADAR1 and ADAR2 both form a homodimer complex. Interestingly, endogenous ADAR3 also appears to form a homodimer complex, indicating the presence of a brain-specific mechanism for ADAR3 dimerization. Homodimer formation may be necessary for ADAR to act as active deaminases. Analysis of dimer complexes consisting of one wild-type and one mutant monomer suggests functional interactions between the two subunits during site-selective RNA editing.