RNA-binding motifs of hnRNP K are critical for induction of antibody diversification by activation-induced cytidine deaminase
RNA-binding motifs of hnRNP K are critical for induction of antibody diversification by activation-induced cytidine deaminase
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DOI：
10.1073/pnas.1921115117
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发表时间：
2020-05
期刊：
影响因子：
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通讯作者：
Ziwei Yin;Maki Kobayashi;Wenjun Hu;Koichi Higashi;N. Begum;K. Kurokawa;T. Honjo
中科院分区：
文献类型：
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作者：
Ziwei Yin;Maki Kobayashi;Wenjun Hu;Koichi Higashi;N. Begum;K. Kurokawa;T. Honjo
意义异质性核糖核蛋白K(HnRNP K)是一种RNA结合蛋白，是激活诱导型胞苷脱氨酶(AID)的辅因子，可诱导免疫球蛋白(Ig)基因的DNA断裂。在这里，我们阐明了GXXG和RGG RNA结合基序对于类开关重组和体细胞超突变是至关重要的。HnRNP K的核定位以及与AID的相互作用也取决于所有的RNA结合基序。这项研究清楚地证明了hnRNP K通过其与RNA的结合能力参与了AID依赖的DNA断裂，提示hnRNP K可能持有并向AID提供一些编辑靶标RNA，最终诱导Ig基因的DNA断裂。激活诱导型胞苷脱氨酶(AID)是类切换重组(CSR)和体细胞超突变(SHM)产生抗体记忆的关键酶。此前，异质性核糖核蛋白K(HnRNP K)被证明是依赖AID的DNA断裂所必需的。在这里，我们分别定义了hnRNP K、GXXGs和RGGs的主要RNA结合基序在K-同源(KH)和K-蛋白质相互作用(KI)结构域中的功能。GXXG和/或RGG突变同样损害CSR、SHM和cMyc/IgH易位，表明这些基序是AID依赖的DNA断裂所必需的。Aid-hnRNP K的相互作用依赖于RNA；因此，这些RNA结合基序的突变如预期的那样取消了与AID的相互作用。一些多嘧啶序列携带典型的hnRNP K结合RNA，参与DNA断裂或修复，以GXXG和RGG基序依赖的方式与hnRNP K结合。GXXG和RGG基序的突变降低了hnRNP K的核保留率，再加上先前发现AID的核定位是其发挥功能所必需的，这些突变体的核保留率较低可能会加剧其功能缺陷，这也是由于它们的RNA结合能力降低所致。总之，hnRNP K通过其所有主要的RNA结合基序促进了AID依赖的DNA断裂。
Significance Heterogeneous nuclear ribonucleoprotein K (hnRNP K), an RNA-binding protein, is the cofactor of activation-induced cytidine deaminase (AID) that induces DNA breaks in immunoglobulin (Ig) genes. Here, we elucidated that the GXXG and RGG RNA-binding motifs were critically necessary for class switch recombination and somatic hypermutation. Nuclear localization of hnRNP K and interaction with AID were also dependent on all of the RNA-binding motifs. This study clearly demonstrated that hnRNP K contributed to AID-dependent DNA breaks through its RNA-binding capacity, suggesting the possibility that hnRNP K holds and presents some editing target RNAs to AID, which eventually induces DNA breaks in Ig genes. Activation-induced cytidine deaminase (AID) is the key enzyme for class switch recombination (CSR) and somatic hypermutation (SHM) to generate antibody memory. Previously, heterogeneous nuclear ribonucleoprotein K (hnRNP K) was shown to be required for AID-dependent DNA breaks. Here, we defined the function of major RNA-binding motifs of hnRNP K, GXXGs and RGGs in the K-homology (KH) and the K-protein-interaction (KI) domains, respectively. Mutation of GXXG, RGG, or both impaired CSR, SHM, and cMyc/IgH translocation equally, showing that these motifs were necessary for AID-dependent DNA breaks. AID–hnRNP K interaction is dependent on RNA; hence, mutation of these RNA-binding motifs abolished the interaction with AID, as expected. Some of the polypyrimidine sequence-carrying prototypical hnRNP K-binding RNAs, which participate in DNA breaks or repair bound to hnRNP K in a GXXG and RGG motif-dependent manner. Mutation of the GXXG and RGG motifs decreased nuclear retention of hnRNP K. Together with the previous finding that nuclear localization of AID is necessary for its function, lower nuclear retention of these mutants may worsen their functional deficiency, which is also caused by their decreased RNA-binding capacity. In summary, hnRNP K contributed to AID-dependent DNA breaks with all of its major RNA-binding motifs.