Yeast Aconitase in Two Locations and Two Metabolic Pathways: Seeing Small Amounts Is Believing

Abstract

The distribution of identical enzymatic activities between different subcellular compartments is a fundamental process of living cells. At present, the Saccharomyces cerevisiae aconitase enzyme has been detected only in mitochondria, where it functions in the tricarboxylic acid (TCA) cycle and is considered a mitochondrial matrix marker. We developed two strategies for physical and functional detection of aconitase in the yeast cytosol: 1) we fused the ␣ peptide of the ␤-galactosidase enzyme to aconitase and observed ␣ complementation in the cytosol; and 2) we created an ACO1-URA3 hybrid gene, which allowed isolation of strains in which the hybrid protein is exclusively targeted to mitochondria. These strains display a specific phenotype consistent with glyoxylate shunt elimination. Together, our data indicate that yeast aconitase isoenzymes distribute between two distinct subcellular compartments and participate in two separate metabolic pathways; the glyoxylate shunt in the cytosol and the TCA cycle in mitochondria. We maintain that such dual distribution phenomena have a wider occurrence than recorded currently, the reason being that in certain cases there is a small fraction of one of the isoenzymes, in one of the locations, making its detection very difficult. We term this phenomenon of highly uneven isoenzyme distribution "eclipsed distribution."

Specialized Function of Yeast Isa1 and Isa2 Proteins in the Maturation of Mitochondrial [4Fe-4S] Proteins

“…5). However, these activities were 10 -15-fold lower than those for expression of native Aco1 in aco1⌬ cells, indicating a comparatively poor maturation of Aco1 in the cytosol (56). Consistent with this idea, Aco1-⌬sp did not rescue the glutamate auxotrophy of aco1⌬ cells (supplemental Fig.…”

“…Further, isa1/2⌬ cells do not elicit signs of a defective core ISC assembly machinery, such as the induction of the yeast iron regulon (22). Maturation of the two tested aconitases, yeast Aco1 and human IRP1, in the yeast cytosol did not involve the Isa proteins and was rather inefficient compared with the process in mitochondria, although it is of physiological importance for the glyoxylate shunt (56). Recent work on the Isa proteins from the protist T. brucei supports the conclusion that the function of the Isa proteins is restricted to the maturation of Fe/S proteins within mitochondria (61).…”

Plant organellar protein targeting: a traffic plan still under construction

“…However, in spite of the apparent absence of a presequence, the protein is localized to the mitochondrial matrix when the cells are enhanced in respiration by growth on the non-fermentable carbon source raffinose [35]. These cellular-responsive dual distributions can be highly disparate in their relative levels, a phenomenon termed eclipsed distributions; this is the case for aconitase in the cytosol and the mitochondrion [36].…”

TCA cycle‐independent acetate metabolism via the glyoxylate cycle in Saccharomyces cerevisiae

“…The cytosolic isozyme Mdh2, but not the peroxisomal enzyme Mdh3, functions in the glyoxylate cycle (Minard and McAlister-Henn, 1991;Van Roermund et al, 1995), whereas the mitochondrial isoform Mdh1 participates in the TCA cycle (McAlister-Henn and Thompson, 1987). Two isoenzymes of ACO encoded by a single gene, ACO1, are distributed between two distinct subcellular compartments, the mitochondria and the cytosol, and participate in the TCA cycle and the glyoxylate cycle, respectively (Gangloff et al, 1990;Regev-Rudzki et al, 2005).…”