Dickson R C, Sheetz R M, Lacy L R
Mol Cell Biol. 1981 Nov;1(11):1048-56. doi: 10.1128/mcb.1.11.1048-1056.1981.
Mutants of Kluyveromyces lactis with elevated uninduced levels of beta-galactosidase (EC 32.1.2.3) activity, constitutive mutants (lac10c), were isolated and characterized to determine the basis for their constitutiveness. These lesions are not operator-type regulatory mutants because they are not closely linked to the beta-galactosidase structural gene. In a constitutive strain having a 7-fold increase in beta-galactosidase activity, the concentration of beta-galactosidase messenger ribonucleic acid (mRNA) was 8- to 10-fold higher than uninduced wild type. The half-life of beta-galactosidase mRNA was the same in the mutant strain (t1/2 = 4.5 +/- 0.2 min) as in uninduced wild-type cells (t1/2 = 4.8 +/- 0.1 min), indicating that the elevated mRNA level in the mutant was not due to a decreased rate of mRNA degradation. Consequently, we hypothesize that the LAC10 product regulates transcription of the beta-galactosidase gene; it probably affects the rate of transcription initiation. Parallel increases in enzyme protein, in constitutive levels of beta-galactosidase activity, and in mRNA further support this position, making translational or posttranslational control by LAC10 unlikely. Several types of data suggest that the LAC10 product functions as a negative regulatory element to prevent transcription. Other data demonstrate that lac10c mutations have pleiotrophic effects, there being constitutive levels not only of beta-galactosidase activity, but also the other lactose-inducible activities of galactokinase (EC 2.7.5.1), galactose-1-phosphate uridyl transferase (EC 2.7.7.10), and lactose transport. It would appear that LAC10 regulates lactose-inducible proteins.
分离并鉴定了具有升高的β-半乳糖苷酶(EC 3.2.1.23)未诱导水平的乳酸克鲁维酵母突变体,即组成型突变体(lac10c),以确定其组成型的基础。这些损伤不是操纵子型调节突变体,因为它们与β-半乳糖苷酶结构基因没有紧密连锁。在β-半乳糖苷酶活性增加7倍的组成型菌株中,β-半乳糖苷酶信使核糖核酸(mRNA)的浓度比未诱导的野生型高8至10倍。β-半乳糖苷酶mRNA在突变菌株中的半衰期(t1/2 = 4.5 +/- 0.2分钟)与未诱导的野生型细胞中的半衰期(t1/2 = 4.8 +/- 0.1分钟)相同,这表明突变体中mRNA水平的升高不是由于mRNA降解速率降低。因此,我们推测LAC10产物调节β-半乳糖苷酶基因的转录;它可能影响转录起始速率。酶蛋白、β-半乳糖苷酶活性组成型水平和mRNA的平行增加进一步支持了这一观点,使得LAC10进行翻译或翻译后控制的可能性不大。几种类型的数据表明,LAC10产物作为一种负调节元件来阻止转录。其他数据表明,lac10c突变具有多效性,不仅β-半乳糖苷酶活性有组成型水平,而且半乳糖激酶(EC 2.7.5.1)、半乳糖-1-磷酸尿苷转移酶(EC 2.7.7.10)和乳糖转运等其他乳糖诱导活性也有组成型水平。看来LAC10调节乳糖诱导蛋白。