PARTICIPATION OF THE TRANSCRIPTION FACTOR Gcn4 IN AMINO ACID HOMEOSTASIS AND AGING IN Saccharomyces cerevisiae

GULIAS, JUAN F.; BERMÚDEZ MORETTI, MARIANA; MUÑOZ, SEBASTIÁN A.; CORREA GARCIA SUSANA

Congreso; SAIB-SAMIGE Joint Meeting 2020; 2020

SAIB y SAMIGE

The transcription factor Gcn4, the global key activator of the GAAC (General Amino acids Control) pathway, maintains amino acid homeostasis in budding yeast by inducing multiple amino acid biosynthetic pathways in response to starvation or imbalance for any amino acid. Gcn4 also induces expression of a variety of genes that mediate purine biosynthesis, organelle biogenesis, endoplasmic reticulum (ER) stress response, mitochondrial carrier proteins and autophagy, while it also represses genes encoding the translation machinery and ribosomes. Amino acids are the building blocks of proteins and, in eukaryotic cells, folding and modifications of membrane and secretory proteins take place in the ER. When the protein folding capacity is exceeded or experimentally impaired, unfolded proteins accumulate in the ER and activate the unfolded protein response (UPR). The cell alleviates the ER stress by the activation of mRNA splicing and the translation of the transcription factor Hac1, which in turn activates the expression of several chaperones such as Kar2. The aim of this work was to study how the loss of amino acid homeostasis affects aging in S. cerevisiae prototrophic cells. It is expected that an addition of amino acids increases the protein synthesis. We measured chronological life span (CLS) and observed that both the presence of all amino acids and the lack of Gcn4 decrease longevity. We also observed that the presence of amino acids triggers KAR2 expression, and that this activation depends on Gcn4. However, we found that the splicing of HAC1 is not altered by the presence of amino acids, and that in gcn4 mutant cells HAC1 splicing is higher than in wild type cells, suggesting that these mutant cells are under ER stress, even in the absence of amino acids or ER chemical stressors. In addition, we analyzed autophagy, a process that produces an increase in the intracellular amino acid pool. We observed that the addition of amino acids reduces the autophagic activity during aging in wild type cells but not in gcn4 cells. Finally, we compared the proteome of wild type and gcn4 mutant cells grown in the absence of amino acids. We found that enzymes for several amino acids biosynthetic pathways were under-expressed in gcn4 cells. In contrast, we found that several proteins related to mitochondrial function, such as enzymes of the TCA cycle, are over-expressed in the mutant strain. This result suggests that in gcn4 cells the synthesis of carbon skeleton precursors of several amino acids may be increased. This could lead to an increment in aerobic respiration that, in turn, could increase oxidative stress as it was reflected by the over-expression of several proteins involved in redox processes. Moreover, we observed in gcn4 cells that there is an over-expression of proteins involved in protein folding. Altogether these results indicate that the loss of amino acid homeostasis caused by the lack of Gcn4 affects longevity in yeast.