Functional Mechanisms Underlying the Antimicrobial Activity of the Oryza sativa Trx-like Protein

Plants are constantly subjected to a variety of environmental stresses and have evolved regulatory responses to overcome unfavorable conditions that might reduce or adversely change a plant’s growth or development. Among these, the regulated production of reactive oxygen species (ROS) as a signaling molecule occurs during plant development and pathogen defense. This study demonstrates the possible antifungal activity of Oryza sativa Tetratricopeptide Domain-containing thioredoxin (OsTDX) protein against various fungal pathogens. The transcription of OsTDX was induced by various environmental stresses known to elicit the generation of ROS in plant cells. OsTDX protein showed potent antifungal activity, with minimum inhibitory concentrations (MICs) against yeast and filamentous fungi ranging between 1.56 and 6.25 and 50 and 100 µg/mL, respectively. The uptake of SYTOX-Green into fungal cells and efflux of calcein from artificial fungus-like liposomes suggest that its killing mechanism involves membrane permeabilization and damage. In addition, irregular blebs and holes apparent on the surfaces of OsTDX-treated fungal cells indicate the membranolytic action of this protein. Our results suggest that the OsTDX protein represents a potentially useful lead for the development of pathogen-resistant plants.

[1]  T. McLellan.  Electrophoresis buffers for polyacrylamide gels at various pH. , 1982, Analytical biochemistry.

[2]  T. Higgins,et al.  A Sensitive, High-Volume, Colorimetric Assay for Protein Phosphatases , 1994, Pharmaceutical Research.

[3]  J. Hamer,et al.  Identification and characterization of Aspergillus nidulans mutants defective in cytokinesis. , 1994, Genetics.

[4]  Karam B. Singh,et al.  Transcription factors in plant defense and stress responses. , 2002, Current opinion in plant biology.

[5]  Seong-Cheol Park,et al.  Antifungal mechanism of a novel antifungal protein from pumpkin rinds against various fungal pathogens. , 2009, Journal of agricultural and food chemistry.

[6]  Jonathan D. G. Jones,et al.  Regulatory Role of SGT1 in Early R Gene-Mediated Plant Defenses , 2002, Science.

[7]  Sang Yeol Lee,et al.  Redox properties of a thioredoxin-like Arabidopsis protein, AtTDX. , 2010, Biochimica et biophysica acta.

[8]  Young Jun Jung,et al.  Functional characterization of thioredoxin h type 5 with antimicrobial activity from Arabidopsis thaliana , 2017, Biotechnology and Bioprocess Engineering.

[9]  D. Shippen,et al.  Dynamic Interactions of Arabidopsis TEN1: Stabilizing Telomeres in Response to Heat Stress , 2016, Plant Cell.

[10]  T. Allen,et al.  Serum-induced leakage of liposome contents. , 1980, Biochimica et biophysica acta.

[11]  M. Esnault,et al.  Ionizing radiation: Advances in plant response , 2010 .

[12]  Seong-Cheol Park,et al.  Hydrophilic Linear Peptide with Histidine and Lysine Residues as a Key Factor Affecting Antifungal Activity , 2018, International journal of molecular sciences.

[13]  Seong-Cheol Park,et al.  Structure-activity relationship of HP (2-20) analog peptide: enhanced antimicrobial activity by N-terminal random coil region deletion. , 2007, Biopolymers.

[14]  H. Askari,et al.  Proteomic responses of rice young panicles to salinity , 2006, Proteomics.

[15]  K. Shinozaki,et al.  Crosstalk in the responses to abiotic and biotic stresses in Arabidopsis: Analysis of gene expression in cytochrome P450 gene superfamily by cDNA microarray , 2004, Plant Molecular Biology.

[16]  K. Sasaki,et al.  A cold‐induced thioredoxin h of rice, OsTrx23, negatively regulates kinase activities of OsMPK3 and OsMPK6 in vitro , 2009, FEBS letters.

[17]  B. Mueller‐Roeber,et al.  A quantitative RT-PCR platform for high-throughput expression profiling of 2500 rice transcription factors , 2007, Plant Methods.

[18]  R. Mittler,et al.  Reactive oxygen gene network of plants. , 2004, Trends in plant science.

[19]  Yi Guo,et al.  An Apoplastic H-Type Thioredoxin Is Involved in the Stress Response through Regulation of the Apoplastic Reactive Oxygen Species in Rice1[C][W][OA] , 2011, Plant Physiology.

[20]  M. Jansen,et al.  The cellular redox state in plant stress biology--a charging concept. , 2010, Plant physiology and biochemistry : PPB.

[21]  Felix Mauch,et al.  The role of abscisic acid in plant-pathogen interactions. , 2005, Current opinion in plant biology.

[22]  P. Urwin,et al.  The interaction of plant biotic and abiotic stresses: from genes to the field. , 2012, Journal of experimental botany.

[23]  Themis Lazaridis,et al.  Antimicrobial peptides bind more strongly to membrane pores. , 2010, Biochimica et biophysica acta.

[24]  Karolina M. Pajerowska-Mukhtar,et al.  Plant Immunity Requires Conformational Charges of NPR1 via S-Nitrosylation and Thioredoxins , 2008, Science.

[25]  K. Shinozaki,et al.  Crosstalk between abiotic and biotic stress responses: a current view from the points of convergence in the stress signaling networks. , 2006, Current opinion in plant biology.

[26]  H. Hirt,et al.  Reactive oxygen species: metabolism, oxidative stress, and signal transduction. , 2004, Annual review of plant biology.

[27]  Seong-Cheol Park,et al.  Deletion of the carboxyl terminal of thioredoxin reductase C of Arabidopsis facilitates oligomerization , 2016, Biotechnology and Bioprocess Engineering.

[28]  K. Kosová,et al.  Plant proteome changes under abiotic stress--contribution of proteomics studies to understanding plant stress response. , 2011, Journal of proteomics.

[29]  N. Gow,et al.  Interactions of fungal pathogens with phagocytes , 2016, Nature Reviews Microbiology.

[30]  S. Wi,et al.  Ultrastructural changes of cell organelles inArabidopsis stems after gamma irradation , 2005, Journal of Plant Biology.

[31]  Seung Joo Cho,et al.  Effect of Leucine and Lysine substitution on the antimicrobial activity and evaluation of the mechanism of the HPA3NT3 analog peptide , 2009, Journal of peptide science : an official publication of the European Peptide Society.

[32]  Seong-Cheol Park,et al.  Amphipathic alpha-helical peptide, HP (2-20), and its analogues derived from Helicobacter pylori: pore formation mechanism in various lipid compositions. , 2008, Biochimica et biophysica acta.

[33]  Kazuo Shinozaki,et al.  Research on plant abiotic stress responses in the post-genome era: past, present and future. , 2010, The Plant journal : for cell and molecular biology.

[34]  Seong-Cheol Park,et al.  Investigation of toroidal pore and oligomerization by melittin using transmission electron microscopy. , 2006, Biochemical and biophysical research communications.

[35]  Hur-Song Chang,et al.  Transcriptional Profiling Reveals Novel Interactions between Wounding, Pathogen, Abiotic Stress, and Hormonal Responses in Arabidopsis1,212 , 2002, Plant Physiology.

[36]  Seong-Cheol Park,et al.  Characterization of a heat-stable protein with antimicrobial activity from Arabidopsis thaliana. , 2007, Biochemical and biophysical research communications.

[37]  Il Ryong Kim,et al.  Molecular mechanism of Arabidopsis thaliana profilins as antifungal proteins. , 2018, Biochimica et biophysica acta. General subjects.

[38]  Hun Heo,et al.  Targeting and synergistic action of an antifungal peptide in an antibiotic drug‐delivery system , 2017, Journal of controlled release : official journal of the Controlled Release Society.

[39]  J. Jorrín-Novo,et al.  Application of proteomics to the assessment of the response to ionising radiation in Arabidopsis thaliana. , 2011, Journal of proteomics.