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Rice ascorbate peroxidase gene family encodes functionally diverse isoforms localized in different subcellular compartments

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Abstract

Aerobic organisms evolved a complex antioxidant system, which protect the cells against oxidative damage caused by partially reduced oxygen intermediates, also known as reactive oxygen species. In plants, ascorbate peroxidases (EC, 1.11.1.11) catalyze the conversion of H2O2 to H2O, using ascorbate as the specific electron donor in this enzymatic reaction. Previously, eight APx genes were identified in the rice (Oryza sativa L.) genome through in silico analysis: two cytosolic isoforms, two putative peroxisomal isoforms, and four putative chloroplastic ones. Using gene-specific probes, we confirmed the presence of the eight APx genes in the rice genome by Southern blot hybridization. Transcript accumulation analysis showed specific expression patterns for each member of the APx family according to developmental stage and in response to salt stress, revealing the complexity of the antioxidant system in plants. Finally, the subcellular localization of rice APx isoforms was determined using GFP-fusion proteins in BY-2 tobacco cells. In agreement with the initial prediction, OSAPX3 was localized in the peroxisomes. On the other hand, the OSAPX6-GFP fusion protein was found in mitochondria of the BY-2 cells, in contrast to the chloroplastic location predicted by sequence analysis. Our findings reveal the functional diversity of the rice APx genes and suggest complementation and coordination of the antioxidant defenses in different cellular compartments during development and abiotic stress.

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Abbreviations

APx:

Ascorbate peroxidase

CaMV:

Cauliflower mosaic virus

CAT:

Catalase

DHAR:

Dehydroascorbate reductase

DIC:

Differential interference contrast

EST:

Expressed sequence tag

GFP:

Green fluorescent protein

GPx:

Glutathione peroxidase

GR:

Glutathione reductase

KO:

Knock-out mutants

MDHAR:

Monodehydroascorbate reductase

PCD:

Programmed cell death

pER:

Peroxisomal endoplasmatic reticulum

Prx:

Peroxidase

ROS:

Reactive oxygen species

SOD:

Superoxide dismutase

SRE:

Splicing regulatory cis-element

TMRE:

Tetramethylrhodamine ethyl ester

3´-UTR:

3´untranslated region

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Acknowledgements

We greatly acknowledge Dr. Peter Waterhouse and Dr. John Watson for critically reading the manuscript. We are grateful to Bernardo Pascarelli and Dr. Radovan Borojevic for helping with the confocal analysis. We thank Dr. Márcio Alves Ferreira, Dr. Gilberto Saccheto Martins, and Dr. Maité Vaslin for helpful discussions and suggestions. We greatly acknowledge the following organizations and researchers for providing us with EST clones, vectors, and antibodies: Dr. Takuji Sasaki, National Institute of Agrobiological Resources-Rice Genome Research Program (NIAS-RGP), Japan; Dr. Susan McCouch, Cornell University, USA; Dr. Mansour Karimi, Ghent University, Belgium; and Dr. Richard Trelease, Arizona State University, USA. This work was supported by FAPERJ, CNPq, and CAPES.

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  1. Larissa Menezes-Benavente

    Present address: Department of Genetics, North Carolina State University, Raleigh, NC, 27695, USA

Authors and Affiliations

  1. Laboratório de Genética Molecular Vegetal, Departamento de Genética, Universidade Federal do Rio de Janeiro, 21944-970, Rio de Janeiro, Brasil

    Felipe Karam Teixeira, Larissa Menezes-Benavente, Vinícius Costa Galvão & Márcia Margis-Pinheiro

  2. Departamento de Bioquímica, Universidade Federal do Rio Grande do Sul, 90035-003, Rio Grande do Sul, Brasil

    Rogério Margis

  3. Laboratório de Genética Vegetal, Departamento de Genética, Universidade Federal do Rio Grande do Sul. Av. Bento Gonçalves, 9500, Prédio 43323, Porto Alegre, RS, Brasil

    Márcia Margis-Pinheiro

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  1. Felipe Karam Teixeira
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Correspondence to Márcia Margis-Pinheiro.

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Teixeira, F.K., Menezes-Benavente, L., Galvão, V.C. et al. Rice ascorbate peroxidase gene family encodes functionally diverse isoforms localized in different subcellular compartments. Planta 224, 300–314 (2006). https://doi.org/10.1007/s00425-005-0214-8

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