Increased frequency of N-region insertion in a murine pre-B-cell line infected with a terminal deoxynucleotidyl transferase retroviral expression vector
Increased frequency of N-region insertion in a murine pre-B-cell line infected with a terminal deoxynucleotidyl transferase retroviral expression vector
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DOI：
10.1128/mcb.7.9.3237-3243.1987
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发表时间：
1987-09
影响因子：
5.3
通讯作者：
N. Landau;D. Schatz;Margaret Rosa;D. Baltimore
中科院分区：
文献类型：
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作者：
N. Landau;D. Schatz;Margaret Rosa;D. Baltimore
研究了末端脱氧核苷酸转移酶（TdT）在免疫球蛋白基因连接位点N区插入中的作用。通过用TdT逆转录病毒表达载体或对照载体感染细胞，获得能够连续重排免疫球蛋白轻链基因且仅在存在或明显不存在TdT方面不同的前B细胞系。然后用基于逆转录病毒的人工免疫球蛋白基因重排底物超感染细胞系。使底物在细胞系中重排，并从细胞系中拯救重排的前病毒。前病毒重排基因的V-J连接的核苷酸序列分析显示，从TdT+细胞系中拯救的前病毒中N-区插入的频率比从TdT-细胞系中拯救的前病毒高5倍，因此在TdT存在下发生的重排中至少有50%具有N区。因此，很明显，TdT可以刺激N区插入，并且该酶可能直接负责在V-J和其他免疫球蛋白和T细胞受体基因连接处添加核苷酸。
The role of terminal deoxynucleotidyl transferase (TdT) in the insertion of N regions into the junctional sites of immunoglobulin genes was investigated. Pre-B-cell lines capable of continuous rearrangement of immunoglobulin light-chain genes and differing only in the presence or apparent absence of TdT were derived by infecting cells with a TdT retroviral expression vector or a control vector. The cell lines were then superinfected with a retrovirus-based artificial immunoglobulin gene rearrangement substrate. The substrate was allowed to rearrange in the cell lines and the rearranged proviruses were rescued from the cell lines. Nucleotide sequence analysis of the V-J junctions of the proviral rearranged genes showed a fivefold greater frequency of N-region insertion in proviruses rescued from the TdT+ cell lines than in those rescued from the TdT- cell lines, so that at least 50% of the rearrangements that occurred in the presence of TdT had N regions. It is thus evident that TdT can stimulate N-region insertion, and the enzyme is presumably directly responsible for adding nucleotides at V-J and other immunoglobulin and T-cell receptor gene junctions.