The mode of action of HC-toxin, a disease determinant of the maize pathogen Cochliobolus carbonum race 1

Hugh A Young, Purdue University

Abstract

Plants are continuously bombarded by an array of potential pathogenic invaders and, therefore, must enlist an equally diverse set of defense response mechanisms to protect themselves. Knowledge of the mechanisms underlying these plant-pathogen interactions is of critical importance in efforts to create disease-resistant crops. Some fungal pathogens rely on the production of toxins to infect their host plants, but, in general, little is known about how such toxins incite disease. Furthermore, mechanisms of host defense in response to these toxins remain elusive. The first part of my thesis focuses on the interaction of Cochliobolus carbonum race 1 (CCR1) and the nonhost plant Arabidopsis thaliana. I seek to elucidate the effects of HC-toxin in Arabidopsis and to provide a greater understanding for the role of this pathotoxin in disease caused by CCR1. Both the root and shoot growth of Arabidopsis were negatively affected when grown on increasing concentrations of HC-toxin. This stunted growth, which is likely in response to the histone deacetylase (HDAC) inhibitor activity of HC-toxin, was reversed when plants were removed from the phytotoxin. Screens using EMS-mutagenized and T-DNA activation-tagged lines resulted in Arabidopsis mutants showing either enhanced sensitivity or greater tolerance to HC-toxin than their wild-type progenitors. Further characterization of the EMS mutant highly sensitive to hc-toxin 4 (hsh4) resulted in phenotypic traits associated with modifications in HDAC machinery. Despite hypersensitivity to HC-toxin, hsh4 did not demonstrate susceptibility to CCR1 infection, suggesting that HDAC inhibition may not be the sole determinant of CCR1 pathogenesis. In addition, mutants isolated from T-DNA activation-tagged lines also provided several gene candidates that may regulate sensitivity to HC-toxin in Arabidopsis. Arabidopsis was also utilized to investigate the role of HC-toxin in susceptibility and defense gene regulation. Exogenous application of HC-toxin induced greater susceptibility to Botrytis cinerea and Alternaria brassicicola. In addition, HC-toxin was able to compromise nonhost resistance in Arabidopsis to CCR1, allowing the pathogen to cause some disease. RNA-blot (Northern) analyses demonstrated that HC-toxin actually induces SA-dependent PR-1 gene expression in Arabidopsis while simultaneously repressing JA-dependent PDF1.2 expression. In addition, HC-toxin induces WRKY70 expression which functions as a regulator between these defense pathways. Therefore, HC-toxin may function as a disease determinant of CCR1 by disrupting the normal regulation of defense response pathways. In the second part of my thesis, I present findings in maize that contradict the current hypothesis for how HC-toxin functions in disease caused by CCR1. Defense genes in maize respond differentially when inoculated with either toxin producing (Tox+) or toxin non-producing (Tox-) isolates of CCR1. Defense genes are rapidly activated and reach a peak before gradually declining in response to Tox- isolates. In contrast, defense genes are not induced appreciably in response to Tox+ isolates, but rather there is a rapid decline of defense gene transcripts after inoculation. Therefore, HC-toxin appears to induce the degradation of defense gene transcripts in maize. In other experiments, Tox+ isolates are able to cause disease on maize lesion mimic mutants that exhibit constitutive defense gene expression. This contradicts the current hypothesis and demonstrates that HC-toxin can induce susceptibility to CCR1 in a manner independent of defense gene suppression. Finally, several HDAC inhibitors other than HC-toxin were ineffective at inducing susceptibility to CCR1, suggesting that HDAC inhibition does not play a critical role in CCR1 pathogenesis.

Degree

Ph.D.

Advisors

Johal, Purdue University.

Subject Area

Plant Pathology|Plant sciences

COinS