Altered Phenotypes in Saccharomyces cerevisiae by Heterologous Expression of Basidiomycete Moniliophthora perniciosa SOD2 Gene
| Autor: | Martin Brendel, Alanna Cibelle Fernandes Pereira, Ana Clara Correia Melgaço, Cristina Pungartnik, Sônia Cristina Oliveira Melo, Regineide Xavier Santos |
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| Rok vydání: | 2015 |
| Předmět: |
Paraquat
diethylnitrosamine super-sensitivity Mutant Saccharomyces cerevisiae Biology Article manganese superoxide dismutase Catalysis Moniliophthora perniciosa Fungal Proteins lcsh:Chemistry Inorganic Chemistry Drug Resistance Bacterial Diethylnitrosamine Physical and Theoretical Chemistry lcsh:QH301-705.5 Molecular Biology Gene Phylogeny Spectroscopy reactive oxygen species Superoxide Dismutase Basidiomycota Genetic Complementation Test Organic Chemistry Wild type Hydrogen Peroxide General Medicine biology.organism_classification Molecular biology Yeast Computer Science Applications Oxidative Stress Coprinopsis cinerea lcsh:Biology (General) lcsh:QD1-999 Mutation functional heterologous expression Heterologous expression |
| Zdroj: | International Journal of Molecular Sciences, Vol 16, Iss 6, Pp 12324-12344 (2015) International Journal of Molecular Sciences Volume 16 Issue 6 Pages 12324-12344 |
| ISSN: | 1422-0067 |
| DOI: | 10.3390/ijms160612324 |
| Popis: | Heterologous expression of a putative manganese superoxide dismutase gene (SOD2) of the basidiomycete Moniliophthora perniciosa complemented the phenotypes of a Saccharomyces cerevisiae sod2Δ mutant. Sequence analysis of the cloned M. perniciosa cDNA revealed an open reading frame (ORF) coding for a 176 amino acid polypeptide with the typical metal-binding motifs of a SOD2 gene, named MpSOD2. Phylogenetic comparison with known manganese superoxide dismutases (MnSODs) located the protein of M. perniciosa (MpSod2p) in a clade with the basidiomycete fungi Coprinopsis cinerea and Laccaria bicolor. Haploid wild-type yeast transformants containing a single copy of MpSOD2 showed increased resistance phenotypes against oxidative stress-inducing hydrogen peroxide and paraquat, but had unaltered phenotype against ultraviolet–C (UVC) radiation. The same transformants exhibited high sensitivity against treatment with the pro-mutagen diethylnitrosamine (DEN) that requires oxidation to become an active mutagen/carcinogen. Absence of MpSOD2 in the yeast sod2Δ mutant led to DEN hyper-resistance while introduction of a single copy of this gene restored the yeast wild-type phenotype. The haploid yeast wild-type transformant containing two SOD2 gene copies, one from M. perniciosa and one from its own, exhibited DEN super-sensitivity. This transformant also showed enhanced growth at 37 °C on the non-fermentable carbon source lactate, indicating functional expression of MpSod2p. The pro-mutagen dihydroethidium (DHE)-based fluorescence assay monitored basal level of yeast cell oxidative stress. Compared to the wild type, the yeast sod2Δ mutant had a much higher level of intrinsic oxidative stress, which was reduced to wild type (WT) level by introduction of one copy of the MpSOD2 gene. Taken together our data indicates functional expression of MpSod2 protein in the yeast S. cerevisiae. |
| Databáze: | OpenAIRE |
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