DIRECT DERIVATION OF CONDITIONALLY IMMORTAL CELL-LINES FROM AN H-2KB-TSA58 TRANSGENIC MOUSE
DIRECT DERIVATION OF CONDITIONALLY IMMORTAL CELL-LINES FROM AN H-2KB-TSA58 TRANSGENIC MOUSE
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DOI：
10.1073/pnas.88.12.5096
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发表时间：
1991-06-01
影响因子：
11.1
通讯作者：
KIOUSSIS, D
中科院分区：
文献类型：
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作者：
JAT, PS;NOBLE, MD;KIOUSSIS, D
对细胞系的研究极大地提高了我们对许多重要生物学问题的理解。通过将永生化癌基因引入感兴趣的细胞类型来促进细胞系的产生。已知在体外使许多不同细胞类型永生化的一种基因编码猿猴病毒40（SV 40）大肿瘤（T）抗原（TAg）。为了避免需要在体外插入基因以产生细胞系，我们创建了携带SV 40 TAg基因的转基因小鼠。由于先前的研究已经表明，在转基因小鼠中的TAg表达与肿瘤发生和异常发展有关，我们利用热不稳定的TAg [来自SV 40株，tsA 58，对转化温度敏感（ts）]来降低体内存在的功能性TAg的水平。为了指导广泛的组织表达，我们使用了小鼠主要组织相容性复合体H-2 K（B）启动子，它既具有广泛的活性，又可以进一步被干扰素诱导。tsA 58 TAg mRNA在携带杂交构建体的所有动物的组织中表达。除胸腺外，所有组织的发育在肉眼下均正常，胸腺始终显示增生。这些小鼠的成纤维细胞和细胞角蛋白+胸腺上皮培养物很容易建立，而不经历危机，并在其生长中有条件地不朽;条件性的程度与检测到的tsA 58 TAg水平相关。H-2K（B）-tsA 58小鼠的一个品系已经通过几代繁殖至纯合性并传递转基因的功能拷贝。
Studies on cell lines have greatly improved our understanding of many important biological questions. Generation of cell lines is facilitated by the introduction of immortalizing oncogenes into cell types of interest. One gene known to immortalize many different cell types in vitro encodes the simian virus 40 (SV40) large tumor (T) antigen (TAg). To circumvent the need for gene insertion in vitro to generate cell lines, we created transgenic mice harboring the SV40 TAg gene. Since previous studies have shown that TAg expression in transgenic mice is associated with tumorigenesis and aberrant development, we utilized a thermolabile TAg [from a SV40 strain, tsA58, temperature sensitive (ts) for transformation] to reduce the levels of functional TAg present in vivo. To direct expression to a broad range of tissues, we used the mouse major histocompatibility complex H-2K(b) promoter, which is both widely active and can be further induced by interferons. tsA58 TAg mRNA was expressed in tissues of all animals harboring the hybrid construct. Development of all tissues was macroscopically normal except for thymus, which consistently showed hyperplasia. Fibroblast and cytokeratin+ thymic epithelial cultures from these mice were readily established without undergoing crisis and were conditionally immortal in their growth; the degree of conditionality was correlated with the levels of tsA58 TAg detected. One strain of H-2K(b)-tsA58 mice has been bred through several generations to homozygosity and transmits a functional copy of the transgene.