Hatanaka Haruyo, Omura Fumihiko, Kodama Yukiko, Ashikari Toshihiko
Suntory Research Center, Osaka 618-8503, Japan.
J Biol Chem. 2009 Jun 5;284(23):15448-57. doi: 10.1074/jbc.M808151200. Epub 2009 Apr 7.
The maltose transporter gene is situated at the MAL locus, which consists of genes for a transporter, maltase, and transcriptional activator. Five unlinked MAL loci (MAL1, MAL2, MAL3, MAL4, and MAL6) constitute a gene family in Saccharomyces cerevisiae. The expression of the maltose transporter is induced by maltose and repressed by glucose. The activity of the maltose transporter is also regulated post-translationally; Mal61p is rapidly internalized from the plasma membrane and degraded by ubiquitin-mediated proteolysis in the presence of glucose. We found that S. cerevisiae strain ATCC20598 harboring MAL21 could grow in maltose supplemented with a non- assimilable glucose analogue, 2-deoxyglucose, whereas strain ATCC96955 harboring MAL61 and strain CB11 with MAL31 and AGT1 could not. These observations implied a Mal21p-specific resistance against glucose-induced degradation. Mal21p found in ATCC20598 has 10 amino acids, including Gly-46 and His-50, that are inconsistent with the corresponding residues in Mal61p. The half-life of Mal21p for glucose-induced degradation was 118 min when expressed using the constitutive TPI1 promoter, which was significantly longer than that of Mal61p (25 min). Studies with mutant cells that are defective in endocytosis or the ubiquitination process indicated that Mal21p was less ubiquitinated than Mal61p, suggesting that Mal21p remains on the plasma membrane because of poor susceptibility to ubiquitination. Mutational studies revealed that both residues Gly-46 and His-50 in Mal21p are essential for the full resistance of maltose transporters against glucose-induced degradation.
麦芽糖转运蛋白基因位于MAL位点，该位点由一个转运蛋白、麦芽糖酶和转录激活因子的基因组成。五个不连锁的MAL位点（MAL1、MAL2、MAL3、MAL4和MAL6）构成了酿酒酵母中的一个基因家族。麦芽糖转运蛋白的表达由麦芽糖诱导，并受到葡萄糖的抑制。麦芽糖转运蛋白的活性也在翻译后受到调节；在葡萄糖存在的情况下，Mal61p会迅速从质膜内化并通过泛素介导的蛋白水解作用降解。我们发现，携带MAL21的酿酒酵母菌株ATCC20598能够在添加了不可同化的葡萄糖类似物2-脱氧葡萄糖的麦芽糖中生长，而携带MAL61的菌株ATCC96955以及携带MAL31和AGT1的菌株CB11则不能。这些观察结果暗示了Mal21p对葡萄糖诱导降解具有特异性抗性。在ATCC20598中发现的Mal21p有10个氨基酸，包括Gly-46和His-50，与Mal61p中的相应残基不一致。当使用组成型TPI1启动子表达时，Mal21p对葡萄糖诱导降解的半衰期为118分钟，明显长于Mal61p的半衰期（25分钟）。对在内吞作用或泛素化过程中存在缺陷的突变细胞的研究表明，Mal21p的泛素化程度低于Mal61p，这表明Mal21p由于对泛素化的敏感性较低而保留在质膜上。突变研究表明，Mal21p中的Gly-46和His-50这两个残基对于麦芽糖转运蛋白对葡萄糖诱导降解的完全抗性都是必不可少的。
