JBC Transcription and Nuclear Factor Monoclonals

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J Biol Chem, Vol. 274, Issue 51, 36729-36733, December 17, 1999

Localization of Nitric-oxide Synthase in Plant Peroxisomes*

Juan B. BarrosoDagger §, Francisco J. Corpas§||, Alfonso CarrerasDagger , Luisa M. Sandalio, R. ValderramaDagger , José M. Palma, José A. Lupiáñez||, and Luis A. del Río

From the  Departamento de Bioquímica, Biología Celular y Molecular de Plantas, Estación Experimental del Zaidín, CSIC, Apartado 419, E-18080 Granada, Spain, the Dagger  Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Experimentales, Universidad de Jaén, Paraje "Las Lagunillas" s/n, E-23071 Jaén, Spain, and the || Departamento de Bioquímica y Biología Molecular, Centro de Ciencias Biológicas, Universidad de Granada, Avenida Fuentenueva s/n, E-18001 Granada, Spain

The presence of nitric-oxide synthase (NOS) in peroxisomes from leaves of pea plants (Pisum sativum L.) was studied. Plant organelles were purified by differential and sucrose density gradient centrifugation. In purified intact peroxisomes a Ca2+-dependent NOS activity of 5.61 nmol of L-[3H]citrulline mg-1 protein min-1 was measured while no activity was detected in mitochondria. The peroxisomal NOS activity was clearly inhibited (60-90%) by different well characterized inhibitors of mammalian NO synthases. The immunoblot analysis of peroxisomes with a polyclonal antibody against the C terminus region of murine iNOS revealed an immunoreactive protein of 130 kDa. Electron microscopy immunogold-labeling confirmed the subcellular localization of NOS in the matrix of peroxisomes as well as in chloroplasts. The presence of NOS in peroxisomes suggests that these oxidative organelles are a cellular source of nitric oxide (NO) and implies new roles for peroxisomes in the cellular signal transduction mechanisms.

* This work was supported by the Dirección General de Enseñanza Superior e Investigación Científica (DGESIC) Grant PB95-0004-01, the Junta de Andalucía (Research Groups CVI 0157 and CVI 0192), Spain, and European Union contract CHRX-CT94-0605.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

§ Contributed equally to the results of this work.

|| To whom correspondence should be addressed: Dept. de Bioquímica, Biología Celular y Molecular de Plantas, Estación Experimental del Zaidín, CSIC, Apartado 419, E-18080 Granada, Spain. Tel.: 34-958-121011; Fax: 34-958-129600; E-mail: javier.corpas@eez.csic.es.

Copyright © 1999 by The American Society for Biochemistry and Molecular Biology, Inc.
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