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We reported previously that the product of the DNA damage-inducible gene of Saccharomyces cerevisiae, DINT, belongs to a family of proteins that are involved in DNA repair and replication. The family includes S. cerevisiae proteins Rad2p and its human homolog XPGC, Rad27p and its mammalian homolog FEN-1, and Exonuclease I (Exo I). Here, we report that Din7p specifically affects metabolism of mitochondrial DNA (mtDNA). We have found that dun1 strains, defective in the transcriptional activation of the DNA damage-inducible genes RNR1, RNR2, and RNR3, exhibit an increased frequency in the formation of the mitochondrial petite ([[Rho].sup.-]) mutants. This high frequency of petites arising in the dun1 strains is significantly reduced by the din7::URA3 allele. On the other hand, overproduction of Din7p from the DIN7 gene placed under control of the GAL1 promoter dramatically increases the frequency of petite formation and the frequency of mitochondrial mutations conferring resistance to erythromycin ([E.sup.r]). The frequencies of chromosomal mutations conferring resistance to canavanine ([Can.sup.r]) or adenine prototrophy ([Ade.sup.+]) are not affected by enhanced synthesis of Din7p. Experiments using Din7p fused to the green fluorescent protein (GFP) and cell fractionation experiments indicate that the protein is located in mitochondria. A possible mechanism that may be responsible for the decreased stability of the mitochondrial genome in S. cerevisiae cells with elevated levels of Din7p is discussed.

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The yeast protein Ccz1p is necessary for vacuolar protein trafficking and biogenesis. In a complex with Mon1p, it mediates fusion of transport intermediates with the vacuole membrane by activating the small GTPase Ypt7p. Additionally, genetic data suggest a role of Ccz1p in earlier transport steps, in the Golgi. In a search for further proteins interacting with Ccz1p, we identified the endosomal soluble N-ethylmaleimide-sensitive factor attachment protein receptor Pep12p as an interaction partner of Ccz1p. Combining the ccz1Δ mutation with deletions of PEP12 or other genes encoding components of the endosomal fusion machinery, VPS21, VPS9 or VPS45, results in synthetic growth phenotypes. The genes MON1 and YPT7 also interact genetically with PEP12. These results suggest that the Ccz1p–Mon1p–Ypt7p complex is involved in fusion of transport vesicles to multiple target membranes in yeast cells. [ABSTRACT FROM AUTHOR]

In Saccharomyces cerevisiae the nicotinic acid moiety of NAD+ can be synthesized from tryptophan using the kynurenine pathway or incorporated directly using nicotinate phosphoribosyl transferase (NPT1). We have identified the genes that encode the enzymes of the kynurenine pathway and for BNA5 (YLR231c) and BNA6 (YFR047c) confirmed that they encode kynureninase and quinolinate phosphoribosyl transferase respectively. We show that deletion of genes encoding kynurenine pathway enzymes are co-lethal with the Δnpt1, demonstrating that no other pathway for the synthesis of nicotinic acid exists in S. cerevisiae. Also, we show that under anaerobic conditions S. cerevisiae is a nicotinic acid auxotroph. [Copyright &y& Elsevier]

The parental strain (AT) of Saccharomyces cerevisiae and mutants, deficient in catalase T (AT), catalase A (AT) or both catalases (AT), grew on ethanol and oleic acid with comparable doubling times. Specific activities of catalase were low in glucose- and ethanol-grown cells. In the two oleic acid-grown A-strains (AT and AT) high catalase activities were found; catalase activity invariably remained low in the AT strain and was never detected in the AT strain. The levels of β-oxidation enzymes in oleic acid-grown cells of the parental and all mutant strains were not significantly different. However, cytochrome C peroxidase activity had increased 8-fold in oleic acid grown A strains (AT and AT) compared to parental strain cells. The degree of peroxisomal proliferation was comparable among the different strains. Catalase A was shown to be located in peroxisomes. Catalase T is most probably cytosolic in nature and/or present in the periplasmic space. [ABSTRACT FROM AUTHOR]

The HEM 12 gene from Saccharomyces cerevisiae encodes uroporphyrinogen decarboxylase which catalyzes the sequential decarboxylation of the four acetyl side chains of uroporphyrinogen to yield coproporphyrinogen, an intermediate in protoheme biosynthesis. The gene was isolated by functional complementation of a hem12 mutant. Sequencing revealed that the HEM12 gene encodes a protein of 362 amino acids with a calculated molecular mass of 41348 Da, The amino acid sequence shares 50% identity with human and rat uroporphyrinogen decarboxylase and shows 40% identity with the N-terminus of an open reading frame described in Synechococcus sp. We determined the sequence of two hem12 mutations which lead to a totally inactive enzyme. They correspond to the amino acid changes Gly33 ↵ Asp and Gly300 ↵ Asp, located in two evolutionarily conserved regions. Each of these substitutions impairs binding of substrates without affecting the overall conformation of the protein. These results argue that a single active center exists in uroporphyrinogen decarboxylase. [ABSTRACT FROM AUTHOR]