Dataset Information

Branched-chain amino acid synthesis is coupled to TOR activation early in the cell cycle in yeast.

ABSTRACT: How cells coordinate their metabolism with division determines the rate of cell proliferation. Dynamic patterns of metabolite synthesis during the cell cycle are unexplored. We report the first isotope tracing analysis in synchronous, growing budding yeast cells. Synthesis of leucine, a branched-chain amino acid (BCAA), increases through the G1 phase of the cell cycle, peaking later during DNA replication. Cells lacking Bat1, a mitochondrial aminotransferase that synthesizes BCAAs, grow slower, are smaller, and are delayed in the G1 phase, phenocopying cells in which the growth-promoting kinase complex TORC1 is moderately inhibited. Loss of Bat1 lowers the levels of BCAAs and reduces TORC1 activity. Exogenous provision of valine and, to a lesser extent, leucine to cells lacking Bat1 promotes cell division. Valine addition also increases TORC1 activity. In wild-type cells, TORC1 activity is dynamic in the cell cycle, starting low in early G1 but increasing later in the cell cycle. These results suggest a link between BCAA synthesis from glucose to TORC1 activation in the G1 phase of the cell cycle.

SUBMITTER: Blank HM

PROVIDER: S-EPMC10481666 | biostudies-literature | 2023 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Branched-chain amino acid synthesis is coupled to TOR activation early in the cell cycle in yeast.

Blank Heidi M HM Reuse Carsten C Schmidt-Hohagen Kerstin K Hammer Staci E SE Hiller Karsten K Polymenis Michael M

EMBO reports 20230727 9

How cells coordinate their metabolism with division determines the rate of cell proliferation. Dynamic patterns of metabolite synthesis during the cell cycle are unexplored. We report the first isotope tracing analysis in synchronous, growing budding yeast cells. Synthesis of leucine, a branched-chain amino acid (BCAA), increases through the G1 phase of the cell cycle, peaking later during DNA replication. Cells lacking Bat1, a mitochondrial aminotransferase that synthesizes BCAAs, grow slower, ...[more]

PMID: 37497662

Similar Datasets

Project description:Branched-chain higher alcohols (BCHAs), or fusel alcohols, including isobutanol, isoamyl alcohol, and active amyl alcohol, are useful compounds in several industries. The yeast Saccharomyces cerevisiae can synthesize these compounds via the metabolic pathways of branched-chain amino acids (BCAAs). Branched-chain amino acid aminotransaminases (BCATs) are the key enzymes for BCHA production via the Ehrlich pathway of BCAAs. BCATs catalyze a bidirectional transamination reaction between branched-chain α-keto acids (BCKAs) and BCAAs. In S. cerevisiae, there are two BCAT isoforms, Bat1 and Bat2, which are encoded by the genes BAT1 and BAT2. Although many studies have shown the effects of deletion or overexpression of BAT1 and BAT2 on BCHA production, there have been no reports on the enhancement of BCHA production by functional variants of BCATs. Here, to improve BCHA productivity, we designed variants of Bat1 and Bat2 with altered enzyme activity by using in silico computational analysis: the Gly333Ser and Gly333Trp Bat1 and corresponding Gly316Ser and Gly316Trp Bat2 variants, respectively. When expressed in S. cerevisiae cells, most of these variants caused a growth defect in minimal medium. Interestingly, the Gly333Trp Bat1 and Gly316Ser Bat2 variants achieved 18.7-fold and 17.4-fold increases in isobutanol above that for the wild-type enzyme, respectively. The enzyme assay revealed that the catalytic activities of all four BCAT variants were lower than that of the wild-type enzyme. Our results indicate that the decreased BCAT activity enhanced BCHA production by reducing BCAA biosynthesis, which occurs via a pathway that directly competes with BCHA production. IMPORTANCE Recently, several studies have attempted to increase the production of branched-chain higher alcohols (BCHAs) in the yeast Saccharomyces cerevisiae. The key enzymes for BCHA biosynthesis in S. cerevisiae are the branched-chain amino acid aminotransaminases (BCATs) Bat1 and Bat2. Deletion or overexpression of the genes encoding BCATs has an impact on the production of BCHAs; however, amino acid substitution variants of Bat1 and Bat2 that could affect enzymatic properties-and ultimately BCHA productivity-have not been fully studied. By using in silico analysis, we designed variants of Bat1 and Bat2 and expressed them in yeast cells. We found that the engineered BCATs decreased catalytic activities and increased BCHA production. Our approach provides new insight into the functions of BCATs and will be useful in the future construction of enzymes optimized for high-level production of BCHAs.

Dataset Information

Branched-chain amino acid synthesis is coupled to TOR activation early in the cell cycle in yeast.

Publications

Branched-chain amino acid synthesis is coupled to TOR activation early in the cell cycle in yeast.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets