Molecular genetics of ICL2, encoding a non-functional isocitrate lyase in saccharomyces cerevisiae

Heinisch, Jürgen J.; Valdés, Eva; Álvarez, José Marcos Alonso; Rodicio, Rosaura

doi:10.1002/(sici)1097-0061(199610)12:13<1285::aid-yea5>3.0.co;2-b

Cited by 15 publications

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References 54 publications

(50 reference statements)

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“…To investigate whether the ICL2 gene encodes a specific 2-methylisocitrate lyase, activities of isocitrate lyase and 2-methylisocitrate lyase were determined in cell extracts of wild-type S. cerevisiae and isogenic icl1⌬, icl2⌬, and icl1⌬ icl2⌬ mutants. All strains were grown on ethanol in shake flask cultures to induce ICL2 transcription (13). Aspartate was used as the sole nitrogen source and as an additional carbon source to circumvent the growth deficiency of icl1 null mutants on ethanol-ammonia media (9,28).…”

Section: Resultsmentioning

confidence: 99%

“…As this pathway is essential for growth on acetate and ethanol, icl1 null mutants are unable to grow on ethanol or acetate (9,28). ICL2 is transcribed in ethanol-grown cultures of wild-type S. cerevisiae, and experiments with ICL2-lacZ fusions indicated that its transcriptional regulation is similar to that of ICL1 (13). However, even the introduction of multiple copies of ICL2 cannot complement the growth deficiency of icl1 null mutants (13).…”

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confidence: 99%

“…ICL2 is transcribed in ethanol-grown cultures of wild-type S. cerevisiae, and experiments with ICL2-lacZ fusions indicated that its transcriptional regulation is similar to that of ICL1 (13). However, even the introduction of multiple copies of ICL2 cannot complement the growth deficiency of icl1 null mutants (13). This indicates that ICL2 does not encode a functional isocitrate lyase.…”

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confidence: 99%

“…This indicates that ICL2 does not encode a functional isocitrate lyase. Since, furthermore, icl2 null mutants have not been found to exhibit a discernible phenotype (13), the physiological function of ICL2 has so far remained an enigma.…”

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confidence: 99%

“…An erroneous assumption about the pathway of propionyl-CoA metabolism in S. cerevisiae has caused some confusion. Based on the assumption that propionyl-CoA metabolism in this yeast involves methyl-malonyl-CoA as an intermediate, results from experiments with 13 C-labeled propionate were interpreted as proof of the occurrence of tight channeling of tricarboxylic acid (TCA) cycle intermediates (30). It was later shown that, instead, the methylcitrate cycle is the key pathway of propionate metabolism (23).…”

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TheSaccharomyces cerevisiae ICL2Gene Encodes a Mitochondrial 2-Methylisocitrate Lyase Involved in Propionyl-Coenzyme A Metabolism

et al. 2000

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The Saccharomyces cerevisiae ICL1 gene encodes isocitrate lyase, an essential enzyme for growth on ethanol and acetate. Previous studies have demonstrated that the highly homologous ICL2 gene (YPR006c) is transcribed during the growth of wild-type cells on ethanol. However, even when multiple copies are introduced, ICL2 cannot complement the growth defect of icl1 null mutants. It has therefore been suggested that ICL2 encodes a nonsense mRNA or nonfunctional protein. In the methylcitrate cycle of propionyl-coenzyme A metabolism, 2-methylisocitrate is converted to succinate and pyruvate, a reaction similar to that catalyzed by isocitrate lyase. To investigate whether ICL2 encodes a specific 2-methylisocitrate lyase, isocitrate lyase and 2-methylisocitrate lyase activities were assayed in cell extracts of wild-type S. cerevisiae and of isogenic icl1, icl2, and icl1 icl2 null mutants. Isocitrate lyase activity was absent in icl1 and icl1 icl2 null mutants, whereas in contrast, 2-methylisocitrate lyase activity was detected in the wild type and single icl mutants but not in the icl1 icl2 mutant. This demonstrated that ICL2 encodes a specific 2-methylisocitrate lyase and that the ICL1-encoded isocitrate lyase exhibits a low but significant activity with 2-methylisocitrate. Subcellular fractionation studies and experiments with an ICL2-green fluorescent protein fusion demonstrated that the ICL2-encoded 2-methylisocitrate lyase is located in the mitochondrial matrix. Similar to that of ICL1, transcription of ICL2 is subject to glucose catabolite repression. In glucose-limited cultures, growth with threonine as a nitrogen source resulted in a ca. threefold induction of ICL2 mRNA levels and of 2-methylisocitrate lyase activity in cell extracts relative to cultures grown with ammonia as the nitrogen source. This is consistent with an involvement of the 2-methylcitrate cycle in threonine catabolism.The complete sequencing of the Saccharomyces cerevisiae genome has yielded a large number of open reading frames with unknown function (11). Some of the newly discovered open reading frames exhibited a strong homology with known yeast genes and were demonstrated to encode hitherto-unknown isoenzymes. An example is the PYK2 gene, which encodes a pyruvate kinase isoenzyme but can restore growth of pyk1 null mutants on glucose only when overexpressed (3). In other cases, the biochemical function of the proteins (if any) encoded by the homologous open reading frames remains unknown.An intriguing case is presented by the ICL2 gene. This gene (YPR006c) exhibits a substantial sequence similarity with ICL1 (13), the unique S. cerevisiae structural gene encoding isocitrate lyase (38% identity at the amino acid level). Isocitrate lyase is a key enzyme of the glyoxylate cycle. As this pathway is essential for growth on acetate and ethanol, icl1 null mutants are unable to grow on ethanol or acetate (9, 28). ICL2 is transcribed in ethanol-grown cultures of wild-type S. cerevisiae, and experiments with ICL2-lacZ fusions indicated that its tra...

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confidence: 99%

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confidence: 99%

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confidence: 99%

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confidence: 99%

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confidence: 99%

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TheSaccharomyces cerevisiae ICL2Gene Encodes a Mitochondrial 2-Methylisocitrate Lyase Involved in Propionyl-Coenzyme A Metabolism

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Investigation of two yeast genes encoding putative isoenzymes of phosphoglycerate mutase

Heinisch

Müller

Schlüter

et al. 1998

Yeast

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Our previous data indicated that GPM1 encodes the only functional phosphoglycerate mutase in yeast. However, in the course of the yeast genome sequencing project, two homologous sequences, designated GPM2 and GPM3, were detected. They have been further investigated in this work. Key residues in the deduced amino acid sequence, shown to be involved in catalysis for Gpm1 (i.e. His8, Arg59, His181) are conserved in both enzymes. Overexpression of the genes under control of their own promoters in a gpm1 deletion mutant did not complement for any of the phenotypes. This could in part be attributed to a lack of expression due to their weak promoters. Higher level expression under the control of the yeast PFK2 promoter partially complemented the gpm1 defects, without restoring detectable enzymatic activity. Nevertheless, deletion of either GPM2 or GPM3, or the two deletions in concert, did not produce any obvious lesions for growth on a variety of different carbon sources, nor did they change the levels of key intermediary metabolites. We conclude that both genes evolved from duplication events and that they probably constitute non‐functional homologues in yeast.

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Transient mRNA responses in chemostat cultures as a method of defining putative regulatory elements: Application to genes involved in Saccharomyces cerevisiae acetyl-coenzyme A metabolism

Berg

Jong-Gubbels

Steensma

1998

Yeast

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Molecular genetics of ICL2, encoding a non-functional isocitrate lyase in saccharomyces cerevisiae

Cited by 15 publications

References 54 publications

TheSaccharomyces cerevisiae ICL2Gene Encodes a Mitochondrial 2-Methylisocitrate Lyase Involved in Propionyl-Coenzyme A Metabolism

TheSaccharomyces cerevisiae ICL2Gene Encodes a Mitochondrial 2-Methylisocitrate Lyase Involved in Propionyl-Coenzyme A Metabolism

Investigation of two yeast genes encoding putative isoenzymes of phosphoglycerate mutase

Transient mRNA responses in chemostat cultures as a method of defining putative regulatory elements: Application to genes involved in Saccharomyces cerevisiae acetyl-coenzyme A metabolism

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