Nitric Oxide Is Critical for Inducing Phosphatidic Acid Accumulation in Xylanase-elicited Tomato Cells

Arjen ten Have,Nicolás Raho,Lorenzo Lamattina,Ana M. Laxalt

doi:10.1074/jbc.m701212200

Abstract

Nitric Oxide (NO) is a second messenger related to development and (a)biotic stress responses in plants. We have studied the role of NO in signaling during plant defense responses upon xylanase elicitation. Treatment of tomato cell cultures with the fungal elicitor xylanase resulted in a rapid and dose-dependent NO accumulation. We have demonstrated that NO is required for the production of the lipid second messenger phosphatidic acid (PA) via the activation of the phospholipase C (PLC) and diacylglycerol kinase (DGK) pathway. Defense-related responses downstream of PA were studied. PA and, correspondingly, xylanase were shown to induce reactive oxygen species production. Scavenging of NO or inhibition of either the PLC or the DGK enzyme diminished xylanase-induced reactive oxygen species production. Xylanase-induced PLDbeta1 and PR1 mRNA levels decreased when NO or PA production were compromised. Finally, we have shown that NO and PA are involved in the induction of cell death by xylanase. Treatment with NO scavenger cPTIO, PLC inhibitor U73122, or DGK inhibitor R59022 diminished xylanase-induced cell death. On the basis of biochemical and pharmacological experimental results, we have shown that PLC/DGK-derived PA represents a novel downstream component of NO signaling cascade during plant defense.

Highlights

One of the second messengers reported to participate in plant defense responses is nitric oxide (NO).2 NO treatments induce plant defense-related transcript accumulation [1, 2], whereas treatments with different inhibitors of NO accumulation compromise the hypersensitive response (HR), a form of programmed cell death induced during plant defense [3]
Tel.: 54-223-4753030; Fax: 54-223-4753150; E-mail: amlaxalt@mdp.edu.ar. 2 The abbreviations used are: NO, nitric oxide; PA, phosphatidic acid; PLC, phospholipase C; PLD, phospholipase D; diacylglycerol kinase (DGK), diacylglycerol (DAG) kinase; induce plant defense-related transcript accumulation [1, 2], whereas treatments with different inhibitors of NO accumulation compromise the hypersensitive response (HR), a form of programmed cell death induced during plant defense [3]
The xylanase-induced PLC/DGK activation required NO (Fig. 3). This is the first evidence showing that NO generates PA via activation of the PLC/DGK pathway

Summary

Introduction

One of the second messengers reported to participate in plant defense responses is nitric oxide (NO).2 NO treatments induce plant defense-related transcript accumulation [1, 2], whereas treatments with different inhibitors of NO accumulation compromise the hypersensitive response (HR), a form of programmed cell death induced during plant defense [3]. In cultured tomato cells treated with xylanase, both PLD and PLC/DGK is activated [24, 29]; in addition, ROS production has been demonstrated [30]. NO and the subsequent PA generation via the PLC/DGK pathway, are required for xylanase-induced ROS production, gene expression, and cell death. These results provide evidence that xylanase-induced NO activates PLC/DGK signaling during plant defense.

Results

Conclusion