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      The protein phosphatase 2A catalytic subunit StPP2Ac2b enhances susceptibility to Phytophthora infestans and senescence in potato

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          Abstract

          The serine/threonine protein phosphatases type 2A (PP2A) are involved in several physiological responses in plants, playing important roles in developmental programs, stress responses and hormone signaling. Six PP2A catalytic subunits (StPP2Ac) were identified in cultivated potato. Transgenic potato plants constitutively overexpressing the catalytic subunit StPP2Ac2b (StPP2Ac2b-OE) were developed to determine its physiological roles. The response of StPP2Ac2b-OE plants to the oomycete Phytophthora infestans, the causal agent of late blight, was evaluated. We found that overexpression of StPP2Ac2b enhances susceptibility to the pathogen. Further bioinformatics, biochemical, and molecular analyses revealed that StPP2Ac2b positively regulates developmental and pathogen-induced senescence, and that P. infestans infection promotes senescence, most likely through induction of StPP2Ac2b expression.

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          A new mathematical model for relative quantification in real-time RT-PCR.

          M. Pfaffl (2001)
          Use of the real-time polymerase chain reaction (PCR) to amplify cDNA products reverse transcribed from mRNA is on the way to becoming a routine tool in molecular biology to study low abundance gene expression. Real-time PCR is easy to perform, provides the necessary accuracy and produces reliable as well as rapid quantification results. But accurate quantification of nucleic acids requires a reproducible methodology and an adequate mathematical model for data analysis. This study enters into the particular topics of the relative quantification in real-time RT-PCR of a target gene transcript in comparison to a reference gene transcript. Therefore, a new mathematical model is presented. The relative expression ratio is calculated only from the real-time PCR efficiencies and the crossing point deviation of an unknown sample versus a control. This model needs no calibration curve. Control levels were included in the model to standardise each reaction run with respect to RNA integrity, sample loading and inter-PCR variations. High accuracy and reproducibility (<2.5% variation) were reached in LightCycler PCR using the established mathematical model.
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            Housekeeping gene selection for real-time RT-PCR normalization in potato during biotic and abiotic stress.

            Nathalie Nicot, Jean-Francois Hausman, Lucien Hoffmann … (2005)
            Plant stress studies are more and more based on gene expression. The analysis of gene expression requires sensitive, precise, and reproducible measurements for specific mRNA sequences. Real-time RT-PCR is at present the most sensitive method for the detection of low abundance mRNA. To avoid bias, real-time RT-PCR is referred to one or several internal control genes, which should not fluctuate during treatments. Here, the non-regulation of seven housekeeping genes (beta-tubulin, cyclophilin, actin, elongation factor 1-alpha (ef1alpha), 18S rRNA, adenine phosphoribosyl transferase (aprt), and cytoplasmic ribosomal protein L2) during biotic (late blight) and abiotic stresses (cold and salt stress) was tested on potato plants using geNorm software. Results from the three experimental conditions indicated that ef1alpha was the most stable among the seven tested. The expression of the other housekeeping genes tested varied upon stress. In parallel, a study of the variability of expression of hsp20.2, shown to be implicated in late blight stress, was realized. The relative quantification of the hsp20.2 gene varied according to the internal control and the number of internal controls used, thus highlighting the importance of the choice of internal controls in such experiments.
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              A translocation signal for delivery of oomycete effector proteins into host plant cells.

              Stephen C Whisson, Petra Boevink, Lucy N. Moleleki … (2007)
              Bacterial, oomycete and fungal plant pathogens establish disease by translocation of effector proteins into host cells, where they may directly manipulate host innate immunity. In bacteria, translocation is through the type III secretion system, but analogous processes for effector delivery are uncharacterized in fungi and oomycetes. Here we report functional analyses of two motifs, RXLR and EER, present in translocated oomycete effectors. We use the Phytophthora infestans RXLR-EER-containing protein Avr3a as a reporter for translocation because it triggers RXLR-EER-independent hypersensitive cell death following recognition within plant cells that contain the R3a resistance protein. We show that Avr3a, with or without RXLR-EER motifs, is secreted from P. infestans biotrophic structures called haustoria, demonstrating that these motifs are not required for targeting to haustoria or for secretion. However, following replacement of Avr3a RXLR-EER motifs with alanine residues, singly or in combination, or with residues KMIK-DDK--representing a change that conserves physicochemical properties of the protein--P. infestans fails to deliver Avr3a or an Avr3a-GUS fusion protein into plant cells, demonstrating that these motifs are required for translocation. We show that RXLR-EER-encoding genes are transcriptionally upregulated during infection. Bioinformatic analysis identifies 425 potential genes encoding secreted RXLR-EER class proteins in the P. infestans genome. Identification of this class of proteins provides unparalleled opportunities to determine how oomycetes manipulate hosts to establish infection.
              Article 0 comments Cited 273 times – based on 0 reviews     Review now
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                Author and article information

                Contributors
                María N. Muñiz García: Role: InvestigationRole: MethodologyRole: Writing – review & editing
                Cecilia Grossi: Role: InvestigationRole: Methodology
                Rita M. Ulloa: Role: Formal analysisRole: ResourcesRole: Writing – review & editing
                Daniela A. Capiati:
                ORCID: https://orcid.org/0000-0002-7169-2003
                Role: ConceptualizationRole: Formal analysisRole: Funding acquisitionRole: Project administrationRole: SupervisionRole: Writing – original draft
                Raffaella Balestrini: Role: Editor
                Journal
                Journal ID (nlm-ta): PLoS One
                Journal ID (iso-abbrev): PLoS One
                Journal ID (publisher-id): plos
                Title: PLoS ONE
                Publisher: Public Library of Science (San Francisco, CA USA )
                ISSN (Electronic): 1932-6203
                Publication date (Electronic): 10 October 2022
                Publication date Collection: 2022
                Volume: 17
                Issue: 10
                Electronic Location Identifier: e0275844
                Affiliations
                [001] Instituto de Investigaciones en Ingeniería Genética y Biología Molecular “Dr. Héctor Torres”, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Buenos Aires, Argentina
                Institute for Sustainable Plant Protection, C.N.R., ITALY
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                https://orcid.org/0000-0002-7169-2003
                Article
                Publisher ID: PONE-D-22-06859
                DOI: 10.1371/journal.pone.0275844
                PMC ID: 9550054
                PubMed ID: 36215282
                SO-VID: 47e25a14-4e54-463f-8f0b-feea8d0b51ec
                Copyright © © 2022 Muñiz García et al
                License:

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                Date received : 7 March 2022
                Date accepted : 25 September 2022
                Page count
                Figures: 6, Tables: 0, Pages: 17
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/501100002923, Consejo Nacional de Investigaciones Científicas y Técnicas;
                Award ID: PIP2015-2017 1220150100415CO
                Award Recipient :
                ORCID: https://orcid.org/0000-0002-7169-2003
                Funded by: funder-id http://dx.doi.org/10.13039/501100003074, Agencia Nacional de Promoción Científica y Tecnológica;
                Award ID: PICT2017-0052
                Award Recipient :
                ORCID: https://orcid.org/0000-0002-7169-2003
                DAC PIP2015-2017 1220150100415CO Consejo Nacional de Investigaciones Científicas y Técnicas https://www.conicet.gov.ar/ DAC PICT2017-0052 Agencia Nacional de Promoción Científica y Tecnológica https://www.argentina.gob.ar/ciencia/agencia The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Subject: Research Article
                Subject: Biology and Life Sciences
                Subject: Plant Science
                Subject: Plant Anatomy
                Subject: Leaves
                Subject: Biology and Life Sciences
                Subject: Cell Biology
                Subject: Cellular Structures and Organelles
                Subject: Chloroplasts
                Subject: Chlorophyll
                Subject: Biology and Life Sciences
                Subject: Cell Biology
                Subject: Plant Cell Biology
                Subject: Chloroplasts
                Subject: Chlorophyll
                Subject: Biology and Life Sciences
                Subject: Plant Science
                Subject: Plant Cell Biology
                Subject: Chloroplasts
                Subject: Chlorophyll
                Subject: Biology and Life Sciences
                Subject: Cell Biology
                Subject: Cellular Types
                Subject: Plant Cells
                Subject: Chloroplasts
                Subject: Chlorophyll
                Subject: Biology and Life Sciences
                Subject: Cell Biology
                Subject: Plant Cell Biology
                Subject: Plant Cells
                Subject: Chloroplasts
                Subject: Chlorophyll
                Subject: Biology and Life Sciences
                Subject: Plant Science
                Subject: Plant Cell Biology
                Subject: Plant Cells
                Subject: Chloroplasts
                Subject: Chlorophyll
                Subject: Physical Sciences
                Subject: Materials Science
                Subject: Materials
                Subject: Pigments
                Subject: Organic Pigments
                Subject: Chlorophyll
                Subject: Biology and Life Sciences
                Subject: Bioengineering
                Subject: Biotechnology
                Subject: Genetic Engineering
                Subject: Genetically Modified Organisms
                Subject: Genetically Modified Plants
                Subject: Engineering and Technology
                Subject: Bioengineering
                Subject: Biotechnology
                Subject: Genetic Engineering
                Subject: Genetically Modified Organisms
                Subject: Genetically Modified Plants
                Subject: Biology and Life Sciences
                Subject: Bioengineering
                Subject: Biotechnology
                Subject: Plant Biotechnology
                Subject: Genetically Modified Plants
                Subject: Engineering and Technology
                Subject: Bioengineering
                Subject: Biotechnology
                Subject: Plant Biotechnology
                Subject: Genetically Modified Plants
                Subject: Biology and Life Sciences
                Subject: Plant Science
                Subject: Plant Biotechnology
                Subject: Genetically Modified Plants
                Subject: Biology and Life Sciences
                Subject: Organisms
                Subject: Eukaryota
                Subject: Plants
                Subject: Genetically Modified Plants
                Subject: Biology and Life Sciences
                Subject: Organisms
                Subject: Eukaryota
                Subject: Plants
                Subject: Solanum
                Subject: Potato
                Subject: Biology and Life Sciences
                Subject: Organisms
                Subject: Eukaryota
                Subject: Plants
                Subject: Vegetables
                Subject: Potato
                Subject: Biology and Life Sciences
                Subject: Plant Science
                Subject: Plant Pathology
                Subject: Plant Pathogens
                Subject: Biology and Life Sciences
                Subject: Genetics
                Subject: Gene Expression
                Subject: Research and Analysis Methods
                Subject: Mathematical and Statistical Techniques
                Subject: Statistical Methods
                Subject: Analysis of Variance
                Subject: Physical Sciences
                Subject: Mathematics
                Subject: Statistics
                Subject: Statistical Methods
                Subject: Analysis of Variance
                Subject: Medicine and Health Sciences
                Subject: Pathology and Laboratory Medicine
                Subject: Pathogens
                Custom metadata
                Data Availability All relevant data are within the manuscript.

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