Saccharomyces cerevisiae contains two functional citrate synthase genes.

Abstract

The tricarboxylic acid cycle occurs within the mitochondria of the yeast Saccharomyces cerevisiae. A nuclear gene encoding the tricarboxylic acid cycle enzyme citrate synthase has previously been isolated (M. Suissa, K. Suda, and G. Schatz, EMBO J. 3:1773-1781, 1984) and is referred to here as CIT1. We report here the isolation, by an immunological method, of a second nuclear gene encoding citrate synthase (CIT2). Disruption of both genes in the yeast genome was necessary to produce classical citrate synthase-deficient phenotypes: glutamate auxotrophy and poor growth on rich medium containing lactate, a nonfermentable carbon source. Therefore, the citrate synthase produced from either gene was sufficient for these metabolic roles. Transcription of both genes was maximally repressed in medium containing both glucose and glutamate. However, transcription of CIT1 but not of CIT2 was derepressed in medium containing a nonfermentable carbon source. The significance of the presence of two genes encoding citrate synthase in S. cerevisiae is discussed.

Suppression of metabolic defects of yeast isocitrate dehydrogenase and aconitase mutants by loss of citrate synthase

“…Also, most mitochondrial protein expression is under global control of the HAP2-5 transcription system but, as mitochondrial function becomes compromised, expression of CIT1, ACO1, and IDH1/2 genes was found to switch to control by the RTG pathway [35], suggesting a cellular need for regulation of glutamate synthesis under this condition. Another indication of a direct correlation is the shared phenotype of glutamate auxotrophy resulting from disruption of individual genes encoding RTG regulatory proteins [36,37], from disruption of ACO1 [38], and from co-disruption of genes encoding major cellular isozymes of citrate synthase [5] or of isocitrate dehydrogenase [25].…”

“…For derivatization, 10 µl samples of clarified supernatants or of standard metabolite mixtures (including TCA cycle intermediates plus aspartate and glutamate at concentrations ranging from 0-2.0 mM) were diluted with 10 µl of water containing labeled internal standards (1.0 mM [1,[5][6][7][8][9][10][11][12][13] Laboratories, respectively) and dried in a microfuge for 3 h using a Speed-Vac. The dry residues were dissolved in 50 µl of 20 mg/ml O-ethylhydroxylamine hydrochloride (Sigma-Aldrich) in pyridine, and reacted for 90 min at 30°C.…”

“…Growth phenotypes associated with disruption of yeast IDH1 and/or IDH2 genes include an inability to grow with acetate as a carbon source, a phenotype shared with several other yeast TCA cycle mutants [5,6], and slow growth with glycerol as a carbon source. The latter phenotype is ameliorated by concomitant disruption of the CIT1 gene encoding the TCA cycle enzyme citrate synthase [7].…”

Subunit structure, expression, and function of NAD(H)-specific isocitrate dehydrogenase in Saccharomyces cerevisiae

“…The S. cerevisiae genes encoding mitochondrial isozymes of citrate synthase (CITI), isolated following enrichment for mRNAs in polysomes bound to mitochondria (41), and of malate dehydrogenase (MDHI), isolated by using immunoscreens of a Agtll expression library (26), were used to construct yeast strains containing chromosomal dismptions in these structural genes. The only dramatic growth phenotype attributable to strains lacking these tricarboxylic acid cycle functions is an inability to grow with acetate as a carbon source (16,26). Based on this observation, we used immunoscreens to identify yeast mutants with defects in NAD(H)-specific isocitrate dehydrogenase in a collection of acetate-negative strains.…”

Multiple bHLH proteins regulate CIT2 expression in Saccharomyces cerevisiae

“…The CIT2 gene encodes citrate synthase, which functions in the TCA and glyoxylate cycles (Kim et al, 1986). The retrograde signalling cascade adjusts carbohydrate and nitrogen metabolism in response to a change in the functional state of the mitochondria (Butow and Avadhani, 2004;Liu and Butow, 2006).…”