Terminal deoxynucleotidyl transferase requires KU80 and XRCC4 to promote N-addition at non-V(D)J chromosomal breaks in non-lymphoid cells
Terminal deoxynucleotidyl transferase requires KU80 and XRCC4 to promote N-addition at non-V(D)J chromosomal breaks in non-lymphoid cells
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DOI：
10.1093/nar/gks585
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发表时间：
2012-09-01
影响因子：
14.9
通讯作者：
Rougeon, Francois
中科院分区：
文献类型：
--
作者：
Boubakour-Azzouz, Imenne;Bertrand, Pascale;Rougeon, Francois
末端脱氧核苷酸转移酶（TdT）是一种DNA聚合酶，通过向V（D）J重组连接处添加非模板核苷酸（N-添加）来增加抗原受体的库。尽管广泛的体外研究TdT的催化活性，合作伙伴的TdT，使N-加成仍有待确定。使用染色体内底物，我们在这里表明，在中国仓鼠卵巢（CHO）细胞中，异位表达TdT有效地促进N-添加在由大范围核酸酶I-SceI产生的染色体双链断裂（DSB）的交界处，N-添加的大小与V（D）J交界处相当。重要的是，在KU 80或XRCC 4缺陷细胞中未观察到N-添加。这些数据表明，在非淋巴细胞的染色体背景下，TdT实际上能够通过严格需要典型非同源末端连接途径KU 80和XRCC 4的组分的过程促进非V（D）J DSB处的N-加成。
Terminal deoxynucleotidyl transferase (TdT) is a DNA polymerase that increases the repertoire of antigen receptors by adding non-templated nucleotides (N-addition) to V(D)J recombination junctions. Despite extensive in vitro studies on TdT catalytic activity, the partners of TdT that enable N-addition remain to be defined. Using an intrachromosomal substrate, we show here that, in Chinese hamter ovary (CHO) cells, ectopic expression of TdT efficiently promotes N-additions at the junction of chromosomal double-strand breaks (DSBs) generated by the meganuclease I-SceI and that the size of the N-additions is comparable with that at V(D)J junctions. Importantly, no N-addition was observed in KU80- or XRCC4-deficient cells. These data show that, in a chromosomal context of non-lymphoid cells, TdT is actually able to promote N-addition at non-V(D)J DSBs, through a process that strictly requires the components of the canonical non-homologous end-joining pathway, KU80 and XRCC4.