The Secreted Antifungal Protein Thionin 2.4 in Arabidopsis thaliana Suppresses the Toxicity of a Fungal Fruit Body Lectin from Fusarium graminearum

Abstract

Plants possess active defense systems and can protect themselves from pathogenic invasion by secretion of a variety of small antimicrobial or antifungal proteins such as thionins. The antibacterial and antifungal properties of thionins are derived from their ability to induce open pore formation on cell membranes of phytopathogens, resulting in release of potassium and calcium ions from the cell. Wheat thionin also accumulates in the cell walls of Fusarium-inoculated plants, suggesting that it may have a role in blocking pathogen infection at the plant cell walls. Here we developed an anti-thionin 2.4 (Thi2.4) antibody and used it to show that Thi2.4 is localized in the cell walls of Arabidopsis and cell membranes of F. graminearum, when flowers are inoculated with F. graminearum. The Thi2.4 protein had an antifungal effect on F. graminearum. Next, we purified the Thi2.4 protein, conjugated it with glutathione-S-transferase (GST) and coupled the proteins to an NHS-activated column. Total protein from F. graminearum was applied to GST-Thi2.4 or Thi2.4-binding columns, and the fungal fruit body lectin (FFBL) of F. graminearum was identified as a Thi2.4-interacting protein. This interaction was confirmed by a yeast two-hybrid analysis. To investigate the biological function of FFBL, we infiltrated the lectin into Arabidopsis leaves and observed that it induced cell death in the leaves. Application of FFBL at the same time as inoculation with F. graminearum significantly enhanced the virulence of the pathogen. By contrast, FFBL-induced host cell death was effectively suppressed in transgenic plants that overexpressed Thi2.4. We found that a 15 kD Thi2.4 protein was specifically expressed in flowers and flower buds and suggest that it acts not only as an antifungal peptide, but also as a suppressor of the FFBL toxicity. Secreted thionin proteins are involved in this dual defense mechanism against pathogen invasion at the plant-pathogen interface.

Transformation of Thionin Genes Using Chitosan Nanoparticle into Potato Plant to Be Resistant to Fungal Infection

“…(12) it was obvious that Thio63 protein cause more inhibition for the radial growth of both A. alternate and R. solani. These results agreed with [16] who worked on the secreted antifungal thionin protein isolated from Arabidopsis thaliana and suppressed the toxicity of fungal wall. …”

“…As to their mode of action, thionins are known to form cation-selective ion channels by binding to phosphatidylserine head groups in lipid bilayer membranes which cause permeabilization and oxidative burst followed by cell death in target cells [49, 50 and 51]. The mechanism also explained simply by [16] as follow: the antibacterial and antifungal properties of thionins are derived from their ability to induce pore formation on cell membranes of phytopathogens, resulting in release of potassium and calcium ions from the cell. …”

“…N-and C-terminal amino acid sequences of these thionins are highly conservative but their nucleotide sequences differentiate to each other [13, 14 and 15]. The mechanism of thionins inhibition to the pathogenic fungi was simply explained by [16] as follow: the antimicrobial properties of thionins are derived from their ability to induce pore formation on cell membranes of phytopathogens, resulting in release of potassium and calcium ions from the cell.…”

Transgenic Potato (Solanumtuberosum) Expressing Two Antifungal Thionin Genes Confer Resistance To Fusarium Spp.

“…While [33] mentioned that the leaf explants were easily injured during the manipulation, which resulted in a low percentage of transformation.Thionin genesisolated from Arabidopsis thaliana were expressed in the transgenic potato cultivars and produced thionin proteins which have the ability to inhibit the fungal growth of both pathogenic fungi (Fusariumsolani and F. oxysporum). These results agreed with [16] who worked on the secreted antifungal thionin protein isolated from Arabidopsis thaliana and suppressed the toxicity of Fusariumgraminearum,also agreed with [34] who concluded that thionins inhibit the growth in vitro of about 20 different fungal plant pathogens including Botrytiscinerea, Fusarium spp., Phytophthorainfestans and Rhizoctoniasolani, [35] also observed that constitutive overexpression of an endogenous thionin in transgenic Arabidopsis resulted in enhanced resistance against Fusarium oxysporum f. sp. matthiolae, which indicates that thionins are defense proteins and [36] were also agreed with our work, who approved that the effective concentration of Thionins of type I, from the endosperms of wheat (Wal, Wa2, Wβ) and barley (Bα, Bβ), and of type II, from barley leaves (BLa, BLb, BLc),giving about 50% inhibition for strains of some bacterial species, such as Clavibactermichiganensis subsp.…”

“…N-and C-terminal amino acid sequences of these thionins are highly conservative but their nucleotide sequences differentiate to each other [13, 14 and 15]. The mechanism of thionins inhibition to the pathogenic fungi was simply explained by [16] as follow: the antimicrobial properties of thionins are derived from their ability to induce pore formation on cell membranes of phytopathogens, resulting in release of potassium and calcium ions from the cell. [17]mentioned that Agrobacterium-mediated transformation has become feasible.…”