Carboxy-terminal domain of AID required for its mRNA complex formation in vivo
Carboxy-terminal domain of AID required for its mRNA complex formation in vivo
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DOI：
10.1073/pnas.0812957106
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发表时间：
2009-02-24
影响因子：
11.1
通讯作者：
Kinoshita, Kazuo
中科院分区：
文献类型：
--
作者：
Nonaka, Taichiro;Doi, Tomomitsu;Kinoshita, Kazuo
激活诱导的胞苷脱氨酶（AID）是IG基因类转换重组（CSR）和体细胞超突变（SHM）的关键。最初，AID被假定为RNA编辑酶，因为它与已知的RNA编辑酶APOBEC 1具有结构同源性。为了支持这一观点，AID与RNA编辑酶具有许多相同的特性，包括核质穿梭和对从头蛋白质合成的依赖性。然而，还没有显示AID是否识别特定的mRNA并编辑它以产生参与CSR或SHM的酶。在这里，我们检查了AID和多聚腺苷酸化[poly（A）（+）] RNA之间的关联在体内，使用UV交联结合poly（A）捕获方法，该方法依赖于生物素化的oligo（dT）和链霉亲和素缀合的珠。我们发现在转染的CH 12细胞中表达的外源性AID和在BL 2细胞中表达的内源性AID都与poly（A）（+）RNA相关。APOBEC 1和APOBEC 3G形成类似的蛋白-poly（A）（+）RNA复合物。然而，所有这些胞苷脱氨酶家族成员，包括AID，与poly（A）（+）RNA的相互作用是间接的。这是对APOBEC 1的预期，已知APOBEC 1通过RNA相互作用辅因子APOBEC 1互补因子（ACF）起作用。此外，AID的羧基末端区域，这是类转换所必需的，也是其与poly（A）（+）RNA相互作用所必需的。这些结果表明，艾滋病的CSR活动需要一个ACF样辅因子，特异性地与艾滋病的羧基末端结构域相互作用。
Activation-induced cytidine deaminase (AID) is essential for the class switch recombination (CSR) and somatic hypermutation (SHM) of Ig genes. Originally, AID was postulated to be an RNA-editing enzyme, because of its structural homology with a known RNA-editing enzyme, APOBEC1. In support of this idea, AID shares many of the properties of RNA-editing enzymes, including nucleocytoplasmic shuttling and a dependency on de novo protein synthesis. However, it has not been shown whether AID recognizes a specific mRNA and edits it to generate an enzyme involved in CSR or SHM. Here, we examined the association between AID and polyadenylated [poly(A)(+)] RNA in vivo, using UV cross-linking coupled with a poly(A) capture method that relies on biotinylated oligo(dT) and streptavidin-conjugated beads. We found that both exogenous AID expressed in transfected CH12 cells and endogenous AID expressed in BL2 cells were associated with poly(A)(+) RNA. Similar protein-poly(A)(+) RNA complexes were formed by APOBEC1 and APOBEC3G. However, the interactions of all of these cytidine deaminase family members, including AID, with poly(A)(+) RNA were indirect. This was expected for APOBEC1, which is known to act through an RNA-interacting cofactor, APOBEC1 complementation factor (ACF). In addition, the carboxy-terminal region of AID, which is essential for class switching, was also required for its interaction with poly(A)(+) RNA. These results suggest that the CSR activity of AID requires an ACF-like cofactor that specifically interacts with the carboxy-terminal domain of AID.