School of Chemical and Biological Engineering, Institute of Chemical Processes, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea.
Theragen Bio Co., Ltd, 145 Gwanggyo-ro, Yeongtong-gu, Suwon-si, Gyeonggi-do 16229, Republic of Korea.
Nucleic Acids Res. 2021 Jan 25;49(2):745-759. doi: 10.1093/nar/gkaa1221.
Gcr1, an important transcription factor for glycolytic genes in Saccharomyces cerevisiae, was recently revealed to have two isoforms, Gcr1U and Gcr1S, produced from un-spliced and spliced transcripts, respectively. In this study, by generating strains expressing only Gcr1U or Gcr1S using the CRISPR/Cas9 system, we elucidate differential activation mechanisms of these two isoforms. The Gcr1U monomer forms an active complex with its coactivator Gcr2 homodimer, whereas Gcr1S acts as a homodimer without Gcr2. The USS domain, 55 residues at the N-terminus existing only in Gcr1U, inhibits dimerization of Gcr1U and even acts in trans to inhibit Gcr1S dimerization. The Gcr1S monomer inhibits the metabolic switch from fermentation to respiration by directly binding to the ALD4 promoter, which can be restored by overexpression of the ALD4 gene, encoding a mitochondrial aldehyde dehydrogenase required for ethanol utilization. Gcr1U and Gcr1S regulate almost the same target genes, but show unique activities depending on growth phase, suggesting that these isoforms play differential roles through separate activation mechanisms depending on environmental conditions.
Gcr1 是酿酒酵母中糖酵解基因的重要转录因子，最近发现它有两种异构体，Gcr1U 和 Gcr1S，分别由未剪接和剪接的转录本产生。在这项研究中，我们通过使用 CRISPR/Cas9 系统生成仅表达 Gcr1U 或 Gcr1S 的菌株，阐明了这两种异构体的差异激活机制。Gcr1U 单体与共激活因子 Gcr2 同源二聚体形成活性复合物，而 Gcr1S 作为同源二聚体而没有 Gcr2。仅存在于 Gcr1U 中的 N 端 55 个残基的 USS 结构域抑制 Gcr1U 的二聚化，甚至以反式作用抑制 Gcr1S 的二聚化。Gcr1S 单体通过直接结合 ALD4 启动子抑制从发酵到呼吸的代谢转换，该转换可以通过过表达编码用于乙醇利用的线粒体醛脱氢酶的 ALD4 基因得到恢复。Gcr1U 和 Gcr1S 调节几乎相同的靶基因，但根据生长阶段表现出独特的活性，表明这些异构体通过独立的激活机制根据环境条件发挥不同的作用。
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