Browsing by Author "Polanco, Ruben"

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    Cloning and functional characterization of the gene encoding the transcription factor Ace1 in the basidiomycete Phanerochaete chrysosporium
    (2006) Polanco, Ruben; Canessa, Paulo; Rivas, Alexis; Larrondo, Luis F.; Lobos, Sergio; Vicuna, Rafael
    In this report we describe the isolation and characterization of a gene encoding the transcription factor Ace 1 (Activation protein of cup 1 Expression) in the white rot fungus Phanerochaete chrysosporium. Pc-ace 1 encodes a predicted protein of 633 amino acids containing the copper-fist DNA binding domain typically found in fungal transcription factors such as Ace 1 Mac 1 and Haa 1 from Saccharomyces cerevisiae. The Pc-ace 1 gene is localized in Scaffold 5, between coordinates 220841 and 222983. A S. cerevisiae ace 1 null mutant strain unable to grow in high-copper medium was fully complemented by transformation with the cDNA of Pc-ace 1. Moreover, Northern blot hybridization studies indicated that Pc-ace 1 cDNA restores copper inducibility of the yeast cup 1 gene, which encodes the metal-binding protein metallothionein implicated in copper resistance. To our knowledge, this is first report describing an Ace 1 transcription factor in basidiomycetes.
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    Expression of genes encoding laccase and manganese-dependent peroxidase in the fungus Ceriporiopsis subvermispora is mediated by an ACE1-like copper-fist transcription factor
    (2009) Miguel Alvarez, Jose; Canessa, Paulo; Mancilla, Rodrigo A.; Polanco, Ruben; Santibanez, Paulina A.; Vicuna, Rafael
    The effect of copper on the expression of genes encoding the ligninolytic enzymes laccase (ICS) and manganese peroxidase (mnp) in Ceriporiopsis subvermispora was evaluated. This metal increased transcript levels of lcs, mnp1 and mnp2. This finding was not unexpected in the case of lcs, since its promoter contains a putative ACE element. Originally characterized in the yeast Saccharomyces cerevisiae, ACE is the target sequence of the ACE1 copper-responsive transcription factor in this microorganism. Analysis of the promoter regions of mnp genes revealed the presence of formerly unnoticed ACE elements. Based on the ace1 gene from Phanerochaete chrysosporium, we isolated and characterized an ACE1-like transcription factor from C. subvermispora (Cs-ACE1) through complementation of a S. cerevisiae ace1 Delta strain. Surprisingly, ACE1 factors from both basidiomycetes exhibit substantial differences, not only structurally but also in their ability to complement the aforementioned yeast strain. Specific binding of Cs-ACE1 to its cognate DNA sequence was confirmed by electrophoretic mobility-shift assays. (C) 2008 Elsevier Inc. All rights reserved.
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    Synthesis, Physicochemical Characterization, and Antimicrobial Evaluation of Halogen-Substituted Non-Metal Pyridine Schiff Bases
    (2024) Carreno, Alexander; Morales-Guevara, Rosaly; Cepeda-Plaza, Marjorie; Paez-Hernandez, Dayan; Preite, Marcelo; Polanco, Ruben; Barrera, Boris; Fuentes, Ignacio; Marchant, Pedro; Fuentes, Juan A.
    Four synthetic Schiff bases (PSB1 [(E)-2-(((4-aminopyridin-3-yl)imino)methyl)-4,6-dibromophenol], PSB2 [(E)-2-(((4-aminopyridin-3-yl)imino)methyl)-4,6-diiodophenol], PSB3 [(E)-2-(((4-aminopyridin-3-yl)imino)methyl)-4-iodophenol], and PSB4 [(E)-2-(((4-aminopyridin-3-yl)imino)methyl)-4-chloro-6-iodophenol]) were fully characterized. These compounds exhibit an intramolecular hydrogen bond between the hydroxyl group of the phenolic ring and the nitrogen of the azomethine group, contributing to their stability. Their antimicrobial activity was evaluated against various Gram-negative and Gram-positive bacteria, and it was found that the synthetic pyridine Schiff bases, as well as their precursors, showed no discernible antimicrobial effect on Gram-negative bacteria, including Salmonella Typhi (and mutant derivatives), Salmonella Typhimurium, Escherichia coli, and Morganella morganii. In contrast, a more pronounced biocidal effect against Gram-positive bacteria was found, including Bacillus subtilis, Streptococcus agalactiae, Streptococcus pyogenes, Enterococcus faecalis, Staphylococcus aureus, and Staphylococcus haemolyticus. Among the tested compounds, PSB1 and PSB2 were identified as the most effective against Gram-positive bacteria, with PSB2 showing the most potent biocidal effects. Although the presence of reactive oxygen species (ROS) was noted after treatment with PSB2, the primary mode of action for PSB2 does not appear to involve ROS generation. This conclusion is supported by the observation that antioxidant treatment with vitamin C only partially mitigated bacterial inhibition, indicating an alternative biocidal mechanism.
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    The Botrytis cinerea Gene Expression Browser
    (2023) Perez-Lara, Gabriel; Moyano, Tomas C.; Vega, Andrea; Larrondo, Luis F.; Polanco, Ruben; Alvarez, Jose M.; Aguayo, Daniel; Canessa, Paulo
    For comprehensive gene expression analyses of the phytopathogenic fungus Botrytis cinerea, which infects a number of plant taxa and is a cause of substantial agricultural losses worldwide, we developed BEB, a web-based B. cinerea gene Expression Browser. This computationally inexpensive web-based application and its associated database contain manually curated RNA-Seq data for B. cinerea. BEB enables expression analyses of genes of interest under different culture conditions by providing publication-ready heatmaps depicting transcript levels, without requiring advanced computational skills. BEB also provides details of each experiment and user-defined gene expression clustering and visualization options. If needed, tables of gene expression values can be downloaded for further exploration, including, for instance, the determination of differentially expressed genes. The BEB implementation is based on open-source computational technologies that can be deployed for other organisms. In this case, the new implementation will be limited only by the number of transcriptomic experiments that are incorporated into the platform. To demonstrate the usability and value of BEB, we analyzed gene expression patterns across different conditions, with a focus on secondary metabolite gene clusters, chromosome-wide gene expression, previously described virulence factors, and reference genes, providing the first comprehensive expression overview of these groups of genes in this relevant fungal phytopathogen. We expect this tool to be broadly useful in B. cinerea research, providing a basis for comparative transcriptomics and candidate gene identification for functional assays.
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    The copper-dependent ACE1 transcription factor activates the transcription of the mco1 gene from the basidiomycete Phanerochaete chrysosporium
    (2008) Canessa, Paulo; Alvarez, Jose Miguel; Polanco, Ruben; Bull, Paulina; Vicuna, Rafael
    We have previously identified and functionally characterized the transcription factor ACE1 (Pc-ACE1) from Phanerochaete chrysosporium. In Saccharomyces cerevisiae, ACE1 activates the copper-dependent transcription of target genes through a DNA sequence element named ACE. However, the possible target gene(s) of Pc-ACE1 were unknown. An in silico search led to the identification of putative ACE elements in the promoter region of mco1, one of the four clustered genes encoding multicopper oxidases (MCOs) in P. chrysosporium. Since copper exerts an effect at the transcriptional level in MCOs from several organisms, in this work we analysed the effect of copper on transcript levels of the clustered MCO genes from P. chrysosporium, with the exception of the transcriptionally inactive mco3. Copper supplementation of cultures produced an increment in transcripts from genes mco1 and mco2, but not from mco4. Electrophoretic mobility-shift assays revealed that Pc-ACE1 binds specifically to a probe containing one of the putative ACE elements found in the promoter of mco1. In addition, using a cell-free transcription system, we showed that in the presence of cuprous ion, Pc-ACE1 induces activation of the promoter of mco1, but not that of mco2.