Function and control of recombination-activating gene activity.
Function and control of recombination-activating gene activity.
复制标题
重组激活基因活性的功能和控制。
DOI：
10.1111/j.1749-6632.1992.tb24626.x
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发表时间：
1992
影响因子：
5.2
通讯作者：
Shinkai,Y
中科院分区：
文献类型：
--
作者：
Alt,FW;Rathbun,G;Oltz,E;Taccioli,G;Shinkai,Y
RAG-1和RAG-2基因协同赋予非淋巴细胞系VDJ重组酶活性。为了明确测试RAG基因的功能，我们建立了缺乏这些基因功能拷贝的小鼠品系。与RAG基因编码VDJ重组酶的组织特异性组分的可能性一致，RAG-2缺陷型小鼠是存活的，但由于不能启动VDJ重组从而产生成熟淋巴细胞而具有严重的联合免疫缺陷。RAG-2缺陷小鼠在淋巴细胞以外的任何组织或谱系中没有明显缺陷，表明VDJ重组酶活性和RAG-2基因功能仅为淋巴细胞发育所需。RAG-1和RAG-2在原代鼠淋巴组织和淋巴骨髓培养物中的表达水平通常远高于转化的前体B细胞系。永久性细胞系中的低水平RAG基因表达是由于具有较低RAG表达水平的细胞的生长导致繁殖期间的下降。在转化的前B细胞系中RAG基因表达的低和可变水平与内源性VDJ重组的低和可变速率相关;因此，这些细胞系不是旨在研究将该活性靶向特定可变区基因区段的机制的实验的可靠模型。为了产生这样一个系统，我们引入RAG基因到B系的热休克诱导型启动子的控制下，热休克处理诱导极高的水平，但短暂的RAG表达伴随着平行诱导VDJ重组酶活性。这些细胞以受调控的方式有效地重排转染的VDJ重组底物，所述受调控的方式依赖于与靶V基因区段相关的转录控制元件的活性。
The RAG-1 and RAG-2 genes synergistically confer VDJ recombinase activity to nonlymphoid cell lines. To unequivocally test RAG gene function, we created lines of mice that lack functional copies of these genes. Consistent with the possibility that RAG gene encode the tissue-specific components of VDJ recombinase, RAG-2-deficient mice are viable but have a severe combined immune deficiency due to inability to initiate VDJ recombination and thereby generate mature lymphocytes. RAG-2-deficient mice have no obvious defect in any tissue or lineage other than lymphocytes, indicating that VDJ recombinase activity and RAG-2-gene function is required only for lymphocyte development. Levels of RAG-1 and RAG-2 expression in primary murine lymphoid tissues and lymphoid bone marrow cultures generally are much higher than those of transformed precursor B-cell lines. Low-level RAG gene expression in permanent cell lines results from a decline during propagation due to outgrowth of cells with lower RAG expression levels. The low and variable level of RAG gene expression in transformed pre-B cell lines correlates with low and variable rates of endogenous VDJ recombination; therefore, such lines are not reliable models for experiments aimed at studying mechanisms that target this activity to particular variable region gene segments. To generate such a system, we introduced RAG genes into B-lineage lines under the control of a heat shock-inducible promoter; heat-shock treatment induces extremely high-level but transient RAG expression accompanied by parallel induction of VDJ recombinase activity. Such cells efficiently rearrange transfected VDJ recombination substrates in a regulated manner that is dependent on the activity of transcriptional control elements associated with the target V gene segments.