Isolation of rat Schwann cell lines: use of SV40 T antigen gene regulated by synthetic metallothionein promoters.
Isolation of rat Schwann cell lines: use of SV40 T antigen gene regulated by synthetic metallothionein promoters.
复制标题
大鼠雪旺细胞系的分离：使用由合成金属硫蛋白启动子调控的 SV40 T 抗原基因。
DOI：
10.1016/0014-4827(89)90037-2
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发表时间：
1989
影响因子：
3.7
通讯作者：
Tennekoon,GI
中科院分区：
文献类型：
--
作者：
Peden,KW;Charles,C;Sanders,L;Tennekoon,GI
Synthetic promoter elements from the mouse metallothionein-I promoter controlling the expression of SV40 T antigen have been tested for their efficacy in cloning rat Schwann cell lines that retained the characteristic properties of these cells and could be passed in culture indefinitely. The constructed promoters contained either four (MT4) or five (MT5) copies of a metal regulatory element 5′ to the CAAT and TATA elements from the HSV-1 thymidine kinase gene. Characterization of these promoters in transient expression assays and transformation assays showed that both MT5and MT4were approximately 10-fold and 15-fold, respectively, weaker than the wild-type MT-I promoter in the presence of heavy metal inducer. However, in the absence of inducer, the basal activity of both MT5and MT4was barely detectable and much lower than that of MT-I. Schwann cells were transfected with plasmids containing the SV40 T antigen gene under the control of the different metallothionein promoters and cell lines were established from each. Only with the MT5and MT4promoters were lines obtained that resembled secondary Schwann cells in culture in their morphology, generation time, and demonstration of contact inhibition. In the presence of zinc, the expression of T antigen in the lines derived with MT5and MT4was about 10-fold lower than that derived with MT-I. On removal of the inducer this level was reduced, and in one cell line T antigen was undetectable. Concomitant with the reduction in T antigen expression there was an increased expression of Po, a protein specific to myelin-forming Schwann cells, and a decreased expression of glial fibrillary acidic protein, a protein expressed only in nonmyelin-forming Schwann cells. These cell lines, therefore, closely resemble untransfected Schwann cells in culture.
