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Tissue-specific localization of aspartic proteinase in developing and germinating barley grains

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Abstract

Resting seeds of several plant species, including barley grains, have been reported to contain aspartic proteinase (EC 3.4.23) activity. Here, the expression of the Hordeum vulgare L. aspartic proteinase (HvAP) was studied in developing and germinating grains by activity measurements as well as by immunocytochemical and in-situ hybridization techniques. Southern blotting suggests the presence of one to two HvAP-encoding genes in the barley genome, while Northern analysis reveals a single 2.1-kb mRNA in grains and vegetative tissues. Western blotting with antibodies to HvAP shows the same subunit structure in different grain parts. In developing grains, HvAP is produced in the embryo, aleurone layer, testa and pericarp, but in the starchy endosperm HvAP is present only in the crushed and depleted area adjacent to the scutellum. During seed maturation, HvAP-encoding mRNA remains in the aleurone layer and in the embryo, but the enzyme disappears from the aleurone cells. The enzyme, however, remains in the degenerating tissues of the testa and pericarp as well as in resting embryo and scutellum. During the first three days of germination, the enzyme reappears in the aleurone layer cells but is not secreted into the starchy endosperm. The HvAP is also expressed in the flowers, stem, leaves, and roots of barley. The wide localization of HvAP in diverse tissues suggests that it may have several functions appropriate to the needs of different tissues.

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Abbreviations

DAA:

days after anthesis

DTT:

dithiothreitol

HvAP:

Hordeum vulgare aspartic proteinase

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Authors and Affiliations

  1. Institute of Biotechnology, University of Helsinki, Karvaamokuja 3, P.O. Box 45, SF-00014, Helsinki, Finland

    Kirsi Törmäkangas, Jukka Kervinen, Anne Östman & Teemu Teeri

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  1. Kirsi Törmäkangas
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  2. Jukka Kervinen
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  3. Anne Östman
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  4. Teemu Teeri
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Correspondence to Kirsi Törmäkangas.

Additional information

Both authors have contributed equally to this work

We thank Mart Saarma, Pia Runeberg-Roos, Alan Schulman and Yrjö Helariutta for helpful discussions during the study, Tiina Arna and Sari Makkonen for their help in proteinase activity experiments as well as Jaana Korhonen (Department of Pathology, University of Helsinki), Salla Marttila and Ilkka Porali (Department of Biology, University of Jyväskylä, Jyväskylä, Finland) for their advice on microscopical techniques. We also thank Liisa Pyhälä and Leena Liesirova for the production of the antibodies to HvAP at the National Public Health Institute, Helsinki. This study was supported by grants from the Ministry of Agriculture and Forestry and the Academy of Finland.

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Törmäkangas, K., Kervinen, J., Östman, A. et al. Tissue-specific localization of aspartic proteinase in developing and germinating barley grains. Planta 195, 116–125 (1994). https://doi.org/10.1007/BF00206299

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