Use of 3-aminotriazole to isolate oleate/acetate non-utilizing mutants of Aspergillus nidulans

Lucas, J. Ramón De; Valenciano, Susana; Amor, Cristina; Laborda, F.

doi:10.4148/1941-4765.1303

Cited by 3 publications

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“…Fatty acid catabolism is another condition that generates H 2 O 2 through peroxisomal ␤-oxidation. In fact, the catalase inhibitor 3-aminotriazole has been used to isolate mutants defective in peroxisome assembly in yeast (45) and A. nidulans (12). Table 1 shows that all mutants grew as the wild type in every condition tested, except in lauric acid and starch plus lauric acid, where the catA/catB double mutant was unable to grow, just as wild-type yeast and A. nidulans strains are unable to grow in lauric acid only when the catalase inhibitor 3-aminotriazole is present (12,45).…”

Section: Vol 179 1997 Regulation Of Catalases In a Nidulans 3287mentioning

confidence: 99%

“…In fact, the catalase inhibitor 3-aminotriazole has been used to isolate mutants defective in peroxisome assembly in yeast (45) and A. nidulans (12). Table 1 shows that all mutants grew as the wild type in every condition tested, except in lauric acid and starch plus lauric acid, where the catA/catB double mutant was unable to grow, just as wild-type yeast and A. nidulans strains are unable to grow in lauric acid only when the catalase inhibitor 3-aminotriazole is present (12,45). In addition to catalases, most aerobic organisms contain peroxidases, and although no A. nidulans peroxidases have been reported, results in Table 1 indicate that unidentified peroxidases can perform part of the catalase function in vivo.…”

Section: Vol 179 1997 Regulation Of Catalases In a Nidulans 3287mentioning

confidence: 99%

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Two divergent catalase genes are differentially regulated during Aspergillus nidulans development and oxidative stress

et al. 1997

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Catalases are ubiquitous hydrogen peroxide-detoxifying enzymes that are central to the cellular antioxidant response. Of two catalase activities detected in the fungus Aspergillus nidulans, the catA gene encodes the spore-specific catalase A (CatA). Here we characterize a second catalase gene, identified after probing a genomic library with catA, and demonstrate that it encodes catalase B. This gene, designated catB, predicts a 721-amino-acid polypeptide (CatB) showing 78% identity to an Aspergillus fumigatus catalase and 61% identity to Aspergillus niger CatR. Notably, similar levels of identity are found when comparing CatB to Escherichia coli catalase HPII (43%), A. nidulans CatA (40%), and the predicted peptide of a presumed catA homolog from A. fumigatus (38%). In contrast, the last two peptides share a 79% identity. The catalase B activity was barely detectable in asexual spores (conidia), disappeared after germination, and started to accumulate 10 h after spore inoculation, throughout growth and conidiation. The catB mRNA was absent from conidia, and its accumulation correlated with catalase activity, suggesting that catB expression is regulated at the transcription level. In contrast, the high CatA activity found in spores was lost gradually during germination and growth. In addition to its developmental regulation, CatB was induced by H 2 O 2 , heat shock, paraquat, or uric acid catabolism but not by osmotic stress. This pattern of regulation and the protective role against H 2 O 2 offered by CatA and CatB, at different stages of the A. nidulans life cycle, suggest that catalase gene redundancy performs the function of satisfying catalase demand at the two different stages of metabolic and genetic regulation represented by growing hyphae versus spores. Alternative H 2 O 2 detoxification pathways in A. nidulans were indicated by the fact that catA/catB double mutants were able to grow in substrates whose catabolism generates H 2 O 2 .

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Section: Vol 179 1997 Regulation Of Catalases In a Nidulans 3287mentioning

confidence: 99%

Section: Vol 179 1997 Regulation Of Catalases In a Nidulans 3287mentioning

confidence: 99%

Two divergent catalase genes are differentially regulated during Aspergillus nidulans development and oxidative stress

et al. 1997

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“…Some fungal mutants unable to grow on acetate are also unable to utilize fatty acids as sole carbon sources (3,27). Selection of A. nidulans mutants unable to grow on fatty acids resulted in the isolation of new alleles of previously identified acu genes (20,41). Fatty acids are first converted to their CoA esters and then, via the cyclic ␤-oxidation pathway, to acetyl-CoA (32,42,69).…”

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confidence: 99%

The facC Gene of Aspergillus nidulans Encodes an Acetate-Inducible Carnitine Acetyltransferase

1998

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Mutations in the facC gene of Aspergillus nidulans result in an inability to use acetate as a sole carbon source. This gene has been cloned by complementation. The proposed translation product of the facC gene has significant similarity to carnitine acetyltransferases (CAT) from other organisms. Total CAT activity was found to be inducible by acetate and fatty acids and repressed by glucose. Acetate-inducible activity was found to be absent in facC mutants, while fatty acid-inducible activity was absent in an acuJ mutant. Acetate induction of facC expression was dependent on the facB regulatory gene, and an expressed FacB fusion protein was demonstrated to bind to 5 facC sequences. Carbon catabolite repression of facC expression was affected by mutations in the creA gene and a CreA fusion protein bound to 5 facC sequences. Mutations in the acuJ gene led to increased acetate induction of facC expression and also of an amdS-lacZ reporter gene, and it is proposed that this results from accumulation of acetate, as well as increased expression of facB. A model is presented in which facC encodes a cytosolic CAT enzyme, while a different CAT enzyme, which is acuJ dependent, is present in peroxisomes and mitochondria, and these activities are required for the movement of acetyl groups between intracellular compartments.

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Characterization of oleate-nonutilizing mutants of Aspergillus nidulans isolated by the 3-amino-1,2,4-triazole positive selection method

Lucas

Valenciano

Domínguez

et al. 1997

Archives of Microbiology

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Conidia of Aspergillus nidulans were mutagenized with ultraviolet light and were incubated on a special selective medium containing the catalase inhibitor 3-amino-1,2,4-triazole. From approximately 5 x 10(7) viable UV-irradiated conidia tested, 423 stable mutants resistant to 3-amino-1,2,4-triazole were recovered, of which 40 were unable to grow on minimal medium with oleic acid as the sole carbon source. These oleate-nonutilizing (Ole-) mutants did not grow on medium with carbon sources requiring functional peroxisomes (oleate, butyrate, acetate, or ethanol), but grew well on medium with carbon sources supposedly not requiring such organelles (glucose, glycerol, l-glutamate, or l-proline). The Ole- mutants carried mutations in one of five nuclear genes affecting acetate utilization: acuJ, acuH, acuE, acuL, and perA. The perA21 strain (DL21) carried a mutation in a gene that is not allelic with any of the known acu loci and displayed a phenotype resembling that described in the Pim- (peroxisome import defective) mutants of Hansenula polymorpha. Hyphae of the perA21 mutant contained a few small peroxisomes with the bulk of peroxisomal enzymes remaining in the 20,000 x g supernatant, but produced wild-type levels of penicillin.

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Use of 3-aminotriazole to isolate oleate/acetate non-utilizing mutants of Aspergillus nidulans

Cited by 3 publications

References 0 publications

Two divergent catalase genes are differentially regulated during Aspergillus nidulans development and oxidative stress

Two divergent catalase genes are differentially regulated during Aspergillus nidulans development and oxidative stress

The facC Gene of Aspergillus nidulans Encodes an Acetate-Inducible Carnitine Acetyltransferase

Characterization of oleate-nonutilizing mutants of Aspergillus nidulans isolated by the 3-amino-1,2,4-triazole positive selection method

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