Habituation to thaxtomin A in hybrid poplar cell suspensions provides enhanced and durable resistance to inhibitors of cellulose synthesis

Abstract

BackgroundThaxtomin A (TA), a phytotoxin produced by the phytopathogen Streptomyces scabies, is essential for the development of potato common scab disease. TA inhibits cellulose synthesis but its actual mode of action is unknown. Addition of TA to hybrid poplar (Populus trichocarpa x Populus deltoides) cell suspensions can activate a cellular program leading to cell death. In contrast, it is possible to habituate hybrid poplar cell cultures to grow in the presence of TA levels that would normally induce cell death. The purpose of this study is to characterize TA-habituated cells and the mechanisms that may be involved in enhancing resistance to TA.ResultsHabituation to TA was performed by adding increasing levels of TA to cell cultures at the time of subculture over a period of 12 months. TA-habituated cells were then cultured in the absence of TA for more than three years. These cells displayed a reduced size and growth compared to control cells and had fragmented vacuoles filled with electron-dense material. Habituation to TA was associated with changes in the cell wall composition, with a reduction in cellulose and an increase in pectin levels. Remarkably, high level of resistance to TA was maintained in TA-habituated cells even after being cultured in the absence of TA. Moreover, these cells exhibited enhanced resistance to two other inhibitors of cellulose biosynthesis, dichlobenil and isoxaben. Analysis of gene expression in TA-habituated cells using an Affymetrix GeneChip Poplar Genome Array revealed that durable resistance to TA is associated with a major and complex reprogramming of gene expression implicating processes such as cell wall synthesis and modification, lignin and flavonoid synthesis, as well as DNA and chromatin modifications.ConclusionsWe have shown that habituation to TA induced durable resistance to the bacterial toxin in poplar cells. TA-habituation also enhanced resistance to two other structurally different inhibitors of cellulose synthesis that were found to target different proteins. Enhanced resistance was associated with major changes in the expression of numerous genes, including some genes that are involved in DNA and chromatin modifications, suggesting that epigenetic changes might be involved in this process.

Enhanced resistance to the cellulose biosynthetic inhibitors, thaxtomin A and isoxaben in Arabidopsis thaliana mutants, also provides specific co-resistance to the auxin transport inhibitor, 1-NPA

“…Better understanding of these complexities will enable a greater understanding of how plant cells respond to toxins and key information about cellulose synthesis [6]. Indeed, similarity of responses expressed in this study particularly by txr1-1 and ixr1-1 , with enhanced multiple resistances to both IXB and NPA suggests that TA and IXB share a common mode of action.…”

“…Testing CBI A. thaliana mutants as presented here or in other plant cell systems such as habituated poplar cell suspensions with resistance to CBIs [6] can aid in identifying common mechanisms across toxins [19,39] and similarities in resistance mechanisms of the mutants. Better understanding of these complexities will enable a greater understanding of how plant cells respond to toxins and key information about cellulose synthesis [6].…”

“…Based on similarity of symptoms produced, TA is believed to be closely related to other cellulose biosynthesis inhibitors (CBIs) such as isoxaben (IXB) and dichlobenil (DCB) [5]. The linkage of the modes of actions of these compounds has been confirmed with habituation studies to TA, revealing cross-resistance to both IXB and DCB [6], although mechanisms of resistance were not investigated.…”

“…The cellular target of TA has not been identified [6,7]. This is in contrast to IXB, where mutant analyses have identified specific cellulose synthase (CesA) complexes (CesA3 and CesA6) from the plasma membrane as toxin targets [8,9].…”

Sensitivity of Aspergillus nidulans to the Cellulose Synthase Inhibitor Dichlobenil: Insights from Wall-Related Genes’ Expression and Ultrastructural Hyphal Morphologies

“…Cell wall inhibitors are a valuable tool to shed light on metabolic pathways regulating extracellular polysaccharide biogenesis and have been indeed used in algae [39]–[43], higher plants [44]–[52], oomycetes [53]–[58] and fungi [59]–[63].…”