Identification by cell fusion of gene sequences that interact with positive trans-acting factors.
Identification by cell fusion of gene sequences that interact with positive trans-acting factors.
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通过细胞融合鉴定与正反式作用因子相互作用的基因序列。
DOI：
10.1126/science.3474782
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发表时间：
1987
期刊：
影响因子：
--
通讯作者：
Bancroft,C
中科院分区：
文献类型：
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作者：
Lufkin,T;Bancroft,C
细胞融合实验已经暗示了特异性内源基因的细胞类型特异性表达中的正或负调节因子。由于不能容易地操纵这些基因，因此无法表征与这些因子相互作用的顺式作用DNA序列。一个转染融合技术，结合稳定的基因转移和形成瞬时异核体，被用来研究这类因子及其DNA结合位点。信使RNA指导的静止，大鼠催乳素启动子区稳定转移到小鼠成纤维细胞后，才检测到大鼠垂体细胞融合，这意味着垂体细胞含有一个积极的细胞类型特异性因子或因子。核连续分析表明融合激活是转录的。融合没有激活稳定转移的大鼠生长激素基因启动子或内源性沉默成纤维细胞催乳素或生长激素基因的表达。通过5′-缺失突变分析，鉴定了大鼠催乳素启动子区细胞融合激活所需的30个碱基对的DNA序列。与以前的结果直接细胞转移该区域的比较意味着，转染融合识别新的调控DNA序列。
Cell fusion experiments have implicated positive or negative regulatory factors in the cell type—specific expression of specialized endogenous genes. The inability to readily manipulate such genes has prevented characterization of the cis-acting DNA sequences that interact with these factors. A transfection-fusion technique, which combined stable gene transfer and formation of transient heterokaryons, was used to study this class of factors and their DNA binding sites. Messenger RNA directed by a quiescent, rat prolactin promoter region stably transferred into mouse fibroblasts was detected only after fusion to rat pituitary cells, implying that pituitary cells contain a positive cell type—specific factor or factors. Nuclear run-on assays showed that fusion activation is transcriptional. Fusion did not activate either a stably transferred rat growth hormone gene promoter or expression of the endogenous silent fibroblast prolactin or growth hormone genes. Analysis by 5′-deletion mutation identified a 30-base pair DNA sequence required for cell fusion activation of the rat prolactin promoter region. Comparison with previous results from direct cellular transfer of this region implies that transfection-fusion identifies novel regulatory DNA sequences.