NADPH Oxidase-Mediated Reactive Oxygen Species Production: Subcellular Localization and Reassessment of Its Role in Plant Defense

Abstract

Chemiluminescence detection of reactive oxygen species (ROS) triggered in tobacco BY-2 cells by the fungal elicitor cryptogein was previously demonstrated to be abolished in cells transformed with an antisense construct of the plasma membrane NADPH oxidase, NtrbohD. Here, using electron microscopy, it has been confirmed that the first hydrogen peroxide production occurring a few minutes after challenge of tobacco cells with cryptogein is plasma membrane located and NtrbohD mediated. Furthermore, the presence of NtrbohD in detergent-resistant membrane fractions could be associated with the presence of NtrbohD-mediated hydrogen peroxide patches along the plasma membrane. Comparison of the subcellular localization of ROS in wild-type tobacco and in plants transformed with antisense constructs of NtrbohD revealed that this enzyme is also responsible for the hydrogen peroxide production occurring at the plasma membrane after infiltration of tobacco leaves with cryptogein. Finally, the reactivity of wild-type and transformed plants to the elicitor and their resistance against the pathogenic oomycete Phytophthora parasitica were examined. NtrbohD-mediated hydrogen peroxide production does not seem determinant for either hypersensitive response development or the establishment of acquired resistance but it is most likely involved in the signaling pathways associated with the protection of the plant cell.

Synergistic Biosynthesis of Biphasic Ethylene and Reactive Oxygen Species in Response to HemibiotrophicPhytophthora parasiticain Tobacco Plants

“…Cryptogein is a 10-kD proteinaceous elicitor purified from culture filtrates of the oomycete P. cryptogea, an avirulent pathogen of tobacco (Ricci et al, 1989), and triggers an HR-like response. Lherminier et al (2009) reported that initial production of H 2 O 2 is mediated by RbohD and occurs within a few minutes after challenge with cryptogein. This event is not sufficient to induce HR development or establish acquired resistance; rather, it is most likely involved in signaling pathways associated with cell protection.…”

Reactive oxygen species and plant resistance to fungal pathogens

“…The cytochemical staining using cerium (III) chloride (CeCl 3 ) is used for a subcellular localisation of H 2 O 2 . The reaction between CeCl 3 and an excess of H 2 O 2 generates electron-dense deposits of cerium perhydroxides that can be observed using transmission electron microscopy (Bestwick et al, 1998;Fester and Hause, 2005;Lherminier et al, 2009;Simon et al, 2013;Xia et al, 2009).…”

Plant Plasma Membrane H+-ATPase in Adaptation of Plants to Abiotic Stresses

“…Nitric oxide (NO) and hydrogen peroxide (H 2 O 2 ) function as signal molecules in plants subjected to abiotic stresses. In plants, the generation of H 2 O 2 seems to be mediated by a plasma membrane bound NADPH oxidase complex (Yang et al, 2007;Lherminier et al, 2009). NO is synthesized via nitric oxide synthase (NOS) and nitrate reductase (Yamasaki & Sakihama, 2000).…”