Study of nitric homeostasis in mitochondrial bioenergetics and in plant defense response to pahogen attack

Author(s)

Halley Caixeta de Oliveira

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia
• Defense date: 2010-03-24

Examining board members

Ione Salgado; Ivan de Godoy Maia; Marcia Regina Braga; Gonçalo Amarante Guimarães Pereira; Paulo Mzzafera

• Advisor: Ione Salgado

Abstract

The control of nitric oxide (NO) homeostasis, determined by a balance between the rate of synthesis and degradation, is essential for its signaling functions. The present study aimed a better understanding of NO homeostasis in plants, focusing on the importance of this molecule in mitochondrial bioenergetics and in plant defense response to pathogen attack. Initially, we carried out a characterization of an NO degradation activity by mitochondria isolated from potato tubers, observing a superoxide-dependent NO consumption, that was stimulated in the presence of NAD(P)H. Assays with different respiratory substrates and inhibitors evidenced the external NAD(P)H dehydrogenases, in addition to complex III, as the main sites of superoxide anion generation for NO consumption. On the other hand, in a comparative analysis with mitochondria isolated from rat liver, a mitochondrial NAD(P)H oxidase activity, non-associated to the respiratory chain, emerged as a superoxide source, in addition to the electron leakage from complexes I and III, for NO consumption. In both cases, the existence of mitochondrial mechanisms of NO degradation was important for the control of its inhibitory effects on respiratory activity. Additionally, the importance of NO synthesis for plant defense was analyzed using the interaction Arabidopsis thaliana- Pseudomonas syringae as a model. Previous works have shown that the nitrate reductase double-deficient mutant of A. thaliana (nia1 nia2) presents reduced NO production and susceptibility to P. syringae, that could result from its impaired nitrogen assimilation. Here, nia1 nia2 plants were cultivated with glutamine or arginine to increase the leaf amino acid content. Despite this, this mutant continued to develop a low NO emission and remained susceptible to bacterial infection, indicating that the susceptibility does not result from reduced amino acid levels. On the other hand, the fumigation of amino acid-recovered nia1 nia2 plants with low concentrations of NO gas reestablished the resistance response. Accordingly, a transcriptomic analysis using DNA microarrays showed that NO treatment induced diverse defense-related genes in infected nia1 nia2 leaves, as those associated to salicylic acid and calcium signaling pathways, pathogenesis-related proteins, cell wall reorganization and synthesis of antimicrobial compounds. Additionally, this analysis indicated new genes as potential targets of NO action, suggesting previously unknown aspects about the role of this signaling molecule in phytopathogenic interactions. In special, we can highlight the possible involvement of NO in the modulation of transcripts related to hormonal signaling in order to allow an attenuating control of certain mechanisms of the defense response. (AU)