Cryptogein-induced initial events in tobacco BY-2 cells: pharmacological characterization of molecular relationship among cytosolic Ca(2+) transients, anion efflux and production of reactive oxygen species.
Ion fluxes and the production of reactive oxygen species (ROS) are early events that follow elicitor treatment or microbial infection. However, molecular mechanisms for these responses as well as their relationship have been controversial and still largely unknown. We here simultaneously monitored the temporal sequence of initial events at the plasma membrane in suspension-cultured tobacco cells (cell line BY-2) in response to a purified proteinaceous elicitor, cryptogein, which induced hypersensitive cell death. The elicitor induced transient rise in cytosolic Ca(2+) concentration ([Ca(2+)](cyt)) showing two distinct peaks, followed by biphasic (rapid/transient and slow/prolonged) Cl(-) efflux and H(+) influx. Pharmacological analyses suggested that the two phases of the [Ca(2+)](cyt) response correspond to Ca(2+) influx through the plasma membrane and an inositol 1,4,5-trisphophate-mediated release of Ca(2+) from intracellular Ca(2+) stores, respectively, and the [Ca(2+)](cyt) transients and the Cl(-) efflux were mutually dependent events regulated by protein phosphorylation. The elicitor also induced production of ROS including (*)O(2)(-) and H(2)O(2), which initiated after the [Ca(2+)](cyt) rise and required Ca(2+) influx, Cl(-) efflux and protein phosphorylation. An inhibitor of NADPH oxidase, diphenylene iodonium, completely inhibited the elicitor-induced production of (*)O(2)(-) and H(2)O(2), but did not affect the [Ca(2+)](cyt) transients. These results suggest that cryptogein-induced plasma membrane Ca(2+) influx is independent of ROS, and NADPH oxidase dependent ROS production is regulated by these series of ion fluxes.
Department of Applied Biological Science, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba, 278-8510 Japan.
Yasuhiro
Y
Kadota
Tatsuaki
T
Goh
Hajime
H
Tomatsu
Ryosuke
R
Tamauchi
Katsumi
K
Higashi
Shoshi
S
Muto
Kazuyuki
K
Kuchitsu
D020418:Algal Proteins / Q000378:metabolism* / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D016923:Cell Death / Q000187:drug effects / Q000502:physiology*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D002712:Chlorides / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005656:Fungal Proteins; D006861:Hydrogen Peroxide / Q000378:metabolism; D007113:Immunity, Innate / Q000502:physiology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007473:Ion Channels / Q000187:drug effects / Q000378:metabolism; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011522:Protons; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D014026:Nicotiana / Q000187:drug effects / Q000378:metabolism*
D020418:Algal Proteins; D002712:Chlorides; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D007473:Ion Channels; D011522:Protons; D017382:Reactive Oxygen Species; C061032:cryptogein protein, Phytophthora cryptogea; D013481:Superoxides; D015544:Inositol 1,4,5-Trisphosphate; D006861:Hydrogen Peroxide; D019255:NADPH Oxidases; D002118:Calcium