Lopez I C, Farrelly F, Butow R A
J Biol Chem. 1981 Jun 25;256(12):6496-501.
We have studied the specification of the apparent molecular weight form of the var1 polypeptide by the var1 determinant region located on mitochondrial DNA of Saccharomyces cerevisiae. A complementation assay has been developed whereby mitochondrial translation products can be labeled selectively in zygotes with 35SO42-. The procedure takes advantage of the fact that met- cys- grande (rho +) strains are unable to incorporate 35SO42- into protein. When such a strain is crossed with a met+ cys petite (rho -) and cells of the mating mixture are labeled with 35SO42- in the presence of cycloheximide under conditions where glycolysis is inoperative, only mitochondrial translation products within zygotes are labeled. Using this procedure with petites containing the var1 determinant region, we find that as much as 75% of the total polypeptide synthesized in zygotes is the form specified by the petite var1 allele. Moreover, mitochondria containing mixed mitochondrial genomes (rho + and rho -) can be isolated from zygotes, and both the rho + and rho - var1 alleles are expressed in vitro. From these results and the analysis of discriminating petites in which var1 recombination can be distinguished from complementation, we conclude that the var1 determinant can specify the apparent molecular weight form of the var1 polypeptide in trans.
我们研究了位于酿酒酵母线粒体DNA上的var1决定簇区域对var1多肽表观分子量形式的特异性。我们开发了一种互补分析方法，通过该方法可以在合子中用35SO42-选择性标记线粒体翻译产物。该方法利用了以下事实：met- cys- grande（rho +）菌株无法将35SO42-掺入蛋白质中。当这样的菌株与met+ cys petite（rho -）杂交，并且在糖酵解不起作用的条件下，在环己酰亚胺存在下用35SO42-标记交配混合物的细胞时，只有合子中的线粒体翻译产物被标记。使用含有var1决定簇区域的小菌落进行此操作，我们发现合子中合成的总多肽中多达75%是由小菌落var1等位基因指定的形式。此外，可以从合子中分离出含有混合线粒体基因组（rho +和rho -）的线粒体，并且rho +和rho - var1等位基因在体外均有表达。根据这些结果以及对可区分var1重组与互补的鉴别小菌落的分析，我们得出结论，var1决定簇可以反式指定var1多肽的表观分子量形式。
