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The Saccharomyces cerevisiae flavodoxin-like proteins Ycp4 and Rfs1 play a role in stress response and in the regulation of genes related to metabolism

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Abstract

SPI1 is a gene whose expression responds to many environmental stimuli, including entry into stationary phase. We have performed a screening to identify genes that activate SPI1 promoter when overexpressed. The phosphatidylinositol-4-phosphate 5-kinase gene MSS4 was identified as a positive activator of SPI1. Another SPI1 transcriptional regulator isolated was the flavodoxin-like gene YCP4. YCP4 and its homolog RFS1 regulate the expression of many genes during the late stages of growth. The double deletion mutant in YCP4 and its homolog RFS1 has an impact on gene expression related to metabolism by increasing the expression of genes involved in hexose transport and glycolysis, and decreasing expression of genes of amino acid metabolism pathways. Genes related to mating and response to pheromone show a decreased expression in the double mutant, while transcription of genes involved in translational elongation is increased. Deletion of these genes, together with the third member of the family, PST2, has a complex effect on the stress response. For instance, double mutant ycp4Δrfs1Δ has an increased response to oxidative stress, but a decreased tolerance to cell-damaging agent SDS. Additionally, this mutation affects chronological aging and slightly increases fermentative capacity.

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Acknowledgments

We thank Chirag Sheth for the critical review of this manuscript. We are indebted to J.C. Igual for the multicopy library. This work has been supported by grants AGL2005-00508 and AGL2008-00060 from the “Ministerio de Educación y Ciencia” to M.O. and A.A. respectively and GRUPOS03/012 from the “Generalitat Valenciana” to both; F.C. was a F.P.U. fellow of the “Ministerio de Educación”.

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  1. Fernando Cardona

    Present address: Unitat de Genètica Molecular, Institut de Biomedicina de València, València, Spain

Authors and Affiliations

  1. Departamento de Biotecnología, Instituto de Agroquímica y Tecnología de Alimentos (CSIC), Av. Agustín Escardino 7, 46980, Paterna, Spain

    Fernando Cardona, Helena Orozco & Agustín Aranda

  2. Departament de Bioquímica i Biologia Molecular, Universitat de València, Burjassot, Spain

    Fernando Cardona, Helena Orozco & Marcel·lí del Olmo

  3. Membrane trafficking and lipid signaling, Génétique Moléculaire, Génomique et Microbiologie, UMR7156 (CNRS-UdS), Strasbourg, France

    Sylvie Friant

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  1. Fernando Cardona
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Correspondence to Agustín Aranda.

Additional information

Communicated by Axel Braghage.

Electronic supplementary material

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203_2011_696_MOESM1_ESM.pdf

Lack of effect of hor2 deletion on SPI1 expression. SPI1 expression tested by northern blot in wild-type strain BY4742 and hor2Δ strain under control conditions (exponential, EXP) and post-diauxic phase (PD) in rich medium. The data shown correspond to the relative amounts of SPI1 mRNA relative to rRNA, and for the same strain transformed with the corresponding empty plasmid. All experiments were carried out in triplicate, and the standard deviation is indicated (PDF 45 kb)

203_2011_696_MOESM2_ESM.pdf

Alignment of flavodoxin-like proteins in S. cerevesiae (A) and other organisms (B). The sequences of Ycp4, Pst2 and Rfs1 of S. cerevisiae (Sc), PbY20 of Paracoccidioides brasiliensis (Pb), Uhp1 of Schyzosaccharomyces pombe (Sp) and Wrba of E. coli (Ec) were taken from NCBI, and the program ClustalX 1.83 was used for the alignment. The last 40 amino acids of Ycp4_Sc are omitted as they were not present in other proteins (PDF 46 kb)

203_2011_696_MOESM3_ESM.pdf

Plasmid loss in mutants of flavodoxin-like genes. Yeast strains transformed with YEPlac357 (A) and the SPI1-lacZ containing plasmids (B) were grown in non-selective medium (YPD), and dilutions were plated on both selective (SC-URA) and non-selective medium (YPD). The percentage of colonies present on the SC-URA plates compared to the YPD plates is represented. Statistically significant data are indicated by *** (p-value<=0.0005) (PDF 46 kb)

203_2011_696_MOESM4_ESM.pdf

Scheme of the results obtained in differential expression between wild-type and mutant ycp4Δrfs1Δ from microarray experiments. Only the genes related to central cellular metabolism are shown. Green text indicates genes differentially expressed at least twofold, and red text indicates those differentially expressed more than twofold in the double mutant compared to wild-type. Relationships with the metabolism of amino acids are shown in blue. The red or green boxes highlight the metabolites whose levels are increased or decreased (respectively) in the double null mutant ycp4Δrfs1Δ. Purple text show up-regulated genes in the ycp4Δ single mutant, whilst red text with an asterisk indicates those overexpressed in both mutants (PDF 95 kb)

203_2011_696_MOESM5_ESM.pdf

Semiquantitative RT–PCR confirmation of selected results obtained by microarray analysis in ycp4Δrfs1Δ strain. A representative result of the replicate analysis carried out with RNA from two independent cultures is shown. ACT1 was used as constitutive gene (PDF 79 kb)

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Cardona, F., Orozco, H., Friant, S. et al. The Saccharomyces cerevisiae flavodoxin-like proteins Ycp4 and Rfs1 play a role in stress response and in the regulation of genes related to metabolism. Arch Microbiol 193, 515–525 (2011). https://doi.org/10.1007/s00203-011-0696-7

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