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Int. J. Mol. Sci. 2015, 16(6), 12324-12344; doi:10.3390/ijms160612324

Altered Phenotypes in Saccharomyces cerevisiae by Heterologous Expression of Basidiomycete Moniliophthora perniciosa SOD2 Gene

1
Departamento de Ciências Biológicas, Laboratório de Biologia de Fungos, Centro de Biotecnologia e Genética, Universidade Estadual de Santa Cruz (UESC), Rodovia Jorge Amado, km 16, Ilhéus, Bahia CEP 45662-900, Brazil
2
Departamento de Ciências Naturais, Universidade Estadual do Sudoeste da Bahia (UESB), Estrada do Bem Querer, km 4, Vitória da Conquista, Bahia CEP 45083-900, Brazil
*
Author to whom correspondence should be addressed.
Academic Editor: Jan Schirawski
Received: 20 December 2014 / Accepted: 8 April 2015 / Published: 1 June 2015
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Abstract

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. View Full-Text
Keywords: reactive oxygen species; manganese superoxide dismutase; functional heterologous expression; diethylnitrosamine super-sensitivity reactive oxygen species; manganese superoxide dismutase; functional heterologous expression; diethylnitrosamine super-sensitivity
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Melo, S.C.; Santos, R.X.; Melgaço, A.C.; Pereira, A.C.F.; Pungartnik, C.; Brendel, M. Altered Phenotypes in Saccharomyces cerevisiae by Heterologous Expression of Basidiomycete Moniliophthora perniciosa SOD2 Gene. Int. J. Mol. Sci. 2015, 16, 12324-12344.

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