C- and N-catabolic utilization of tricarboxylic acid cycle-related amino acids by Scheffersomyces stipitis and other yeasts.

Scheffersomyces stipitis and the closely related yeast Candida shehatae assimilated the L-amino acids glutamate, aspartate and proline as both carbon and nitrogen sole sources. We also found this rarely investigated ability in ascomycetous species such as Candida glabrata, C. reukaufii, C. utilis, Debaryomyces hansenii, Kluyveromyces lactis, K. marxianus, Candida albicans, L. elongisporus, Meyerozyma guilliermondii, C. maltosa, Pichia capsulata and Yarrowia lipolytica and in basidiomycetous species such as Rhodotorula rubra and Trichosporon beigelii. Glutamate was a very efficient carbon source for Sc. stipitis, which enabled a high biomass yield/mole, although the growth rate was lower when compared to growth on glucose medium. The cells secreted waste ammonium during growth on glutamate alone. In Sc. stipitis cultures grown in glucose medium containing glutamate as the nitrogen source the biomass yield was maximal, and ethanol concentration and specific ethanol formation rate were significantly higher than in glucose medium containing ammonium as the nitrogen source. Mainly C-assimilation of glutamate but also N-assimilation in glucose-containing medium correlated with enhanced activity of the NAD-dependent glutamate dehydrogenase 2 (GDH2). A Δgdh2 disruptant was unable to utilize glutamate as either a carbon or a nitrogen source; moreover, this disruptant was also unable to utilize aspartate as a carbon source. The mutation was complemented by retransformation of the GDH2 ORF into the Δgdh2 strain. The results show that Gdh2p plays a dual role in Sc. stipitis as both C- and N-catabolic enzyme, which indicates its role as an interface between the carbon and nitrogen metabolism of this yeast.

The 3' ends of mature transcripts are generated by a processosome complex in fission yeast mitochondria.

In this article, we report on the genetic analysis of the Schizosaccharomyces pombe open reading frames SPCC1322.01 and SPAC637.11, respectively, which encode proteins that are similar to the exoribonuclease Dss1p and the RNA helicase Suv3p, respectively, forming the mitochondrial degradosome of Saccharomyces cerevisiae. While the helicase Suv3p is exchangeable between S. cerevisiae and S. pombe, the functions of Dss1p and the putative fission yeast RNase protein are specific for each species. Unlike S. cerevisiae mutants lacking a functional degradosome, the major defect of fission yeast knock-out strains is their inability to perform downstream processing of transcripts. In addition, the lack of pah1 results in instability of mitochondrial RNA ends. Overexpression of par1 and pah1 has no significant effect on the steady-state levels of mitochondrial RNAs. The Pet127p-stimulated RNA degradation activity is independent of Par1p/Pah1p in fission yeast mitochondria. The results presented herein indicate that both fission yeast proteins play only a minor role (if at all) in mitochondrial RNA degradation. We assume that the RNA-degrading function was taken over by other enzymes in fission yeast mitochondria, while the former degradosome proteins were recruited to new cellular pathways, for example, RNA processing in fission yeast (as discussed in this article) or mitochondrial DNA replication, apoptosis, or chromatin maintenance in eukaryotes, during evolution.

RNA degradation in fission yeast mitochondria is stimulated by a member of a new family of proteins that are conserved in lower eukaryotes.

We report here on the role of open reading frame (ORF) SPCC1183.04c of Schizosaccharomyces pombe in mitochondrial RNA metabolism. A mutant deleted for this ORF on chromosome III accumulates mitochondrial transcripts with the exception of the cob mRNA. A detailed Northern blot analysis showed that the effect results from a decrease in RNA degradation but not from RNA processing deficiencies. Overexpression of the SPCC1183.04c gene in a S. pombe wild-type strain is characterized by slow growth at 37 degrees C on non-fermentable carbon sources and a significant reduction of steady-state levels of mitochondrial transcripts. A NCBI BLASTP search with the amino acid sequence deduced from the S. pombe gene identified significant similarity to a number of proteins in fungi (e.g. Ascomycota, Basidiomycota) and in some non-fungal eukaryotes (e.g. ciliate, slime mold, red algae). By heterologous expression of SPCC1183.04c in a Saccharomyces cerevisiae pet127Delta strain, we demonstrate that the fission yeast protein and Pet127p from S. cerevisiae function similarly: The fission yeast gene complemented the respiratory defect associated with the pet127Delta allele and partially restored the RNA processing phenotype. Although it lacks any recognizable targeting signal, the S. pombe protein is imported into S. cerevisiae mitochondria in vivo. We conclude from our results that the fission yeast SPCC1183.04c gene is a member of a new protein family that functions to stimulate mitochondrial RNA degradation, a function that is conserved within the mitochondria of lower eukaryotes but seems to have been replaced by alternative pathways in metazoans and higher plants.