May G S, Waring R B, Morris N R
Department of Pharmacology, UMDNJ, R.W. Johnson Medical School, Piscataway.
Cell Motil Cytoskeleton. 1990;16(3):214-20. doi: 10.1002/cm.970160308.
We have constructed a chimeric beta-tubulin gene that places the structural gene for the tubC beta-tubulin of Aspergillus nidulans under the control of the benA beta-tubulin promoter. Introduction of either this chimeric gene or a second wild-type benA gene into a benomyl-resistant benA22 strain causes it to become benomyl sensitive, indicating that the introduced genes are functional. Analysis of the tubulin proteins synthesized in benA22 strains into which a second wild-type benA beta-tubulin gene was transformed showed that the total amount of beta-tubulin protein was the same as in the parental strain with a single benA gene. Thus the level of beta-tubulin must be regulated. This was also true of transformants carrying an extra copy of the chimeric beta-tubulin gene. The total amount of beta-tubulin was the same as in the parental strain. Two-dimensional gel analysis showed that the endogenous benA22 and the introduced chimeric tubC gene contributed equally to the total beta-tubulin pool. The fact that one-half of the benA beta-tubulin could be replaced by tubC beta-tubulin with no effect on the growth of the cells suggests that the benA and tubC beta-tubulins are functionally interchangeable.
我们构建了一个嵌合β-微管蛋白基因，该基因将构巢曲霉tubCβ-微管蛋白的结构基因置于benAβ-微管蛋白启动子的控制之下。将这个嵌合基因或第二个野生型benA基因导入对苯菌灵有抗性的benA22菌株中，会使其对苯菌灵变得敏感，这表明导入的基因是有功能的。对转入第二个野生型benAβ-微管蛋白基因的benA22菌株中合成的微管蛋白进行分析表明，β-微管蛋白的总量与含有单个benA基因的亲本菌株相同。因此，β-微管蛋白的水平必定受到调控。携带嵌合β-微管蛋白基因额外拷贝的转化体也是如此。β-微管蛋白的总量与亲本菌株相同。二维凝胶分析表明，内源性benA22和导入的嵌合tubC基因对总的β-微管蛋白库的贡献相同。benAβ-微管蛋白的一半可以被tubCβ-微管蛋白替代而对细胞生长没有影响，这一事实表明benA和tubCβ-微管蛋白在功能上是可互换的。
