H₂O₂ Intensifies CN^--Induced Apoptosis in Pea Leaves

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V. D. Samuilov^*, D. B. Kiselevsky, S. V. Sinitsyn, A. A. Shestak, E. M. Lagunova, and A. V. Nesov

Department of Physiology of Microorganisms, Biological Faculty, Lomonosov Moscow State University, 119992 Moscow, Russia; fax: (495) 939-3807; E-mail: vdsamuilov@mail.ru

^* To whom correspondence should be addressed.

Received October 11, 2005; Revision received November 16, 2005

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H₂O₂ intensifies CN^--induced apoptosis in stoma guard cells and to lesser degree in basic epidermal cells in peels of the lower epidermis isolated from pea leaves. The maximum effect of H₂O₂ on guard cells was observed at 10^-4 M. By switching on non-cyclic electron transfer in chloroplasts menadione and methyl viologen intensified H₂O₂ generation in the light, but prevented the CN^--induced apoptosis in guard cells. The light stimulation of CN^- effect on guard cell apoptosis cannot be caused by disturbance of the ribulose-1,5-bisphosphate carboxylase function and associated OH^* generation in chloroplasts with participation of free transition metals in the Fenton or Haber-Weiss type reactions as well as with participation of the FeS clusters of the electron acceptor side of Photosystem I. Menadione and methyl viologen did not suppress the CN^--induced apoptosis in epidermal cells that, unlike guard cells, contain mitochondria only, but not chloroplasts. Quinacrine and diphenylene iodonium, inhibitors of NAD(P)H oxidase of cell plasma membrane, had no effect on the respiration and photosynthetic O₂ evolution by leaf slices, but prevented the CN^--induced guard cell death. The data suggest that NAD(P)H oxidase of guard cell plasma membrane is a source of reactive oxygen species (ROS) needed for execution of CN^--induced programmed cell death. Chloroplasts and mitochondria were inefficient as ROS sources in the programmed death of guard cells. When ROS generation is insufficient, exogenous H₂O₂ exhibits a stimulating effect on programmed cell death. H₂O₂ decreased the inhibitory effects of DCMU and DNP-INT on the CN^--induced apoptosis of guard cells. Quinacrine, DCMU, and DNP-INT had no effect on CN^--induced death of epidermal cells.

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KEY WORDS: apoptosis, reactive oxygen species, chloroplasts, mitochondria, plasma membrane, Pisum sativum L.

DOI: 10.1134/S0006297906040067

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