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Knockdown of the 7S globulin subunits shifts distribution of nitrogen sources to the residual protein fraction in transgenic soybean seeds

Plant Cell Reports volume 33, pages 1963–1976 (2014)Cite this article

Abstract

Key message

A platform of gene silencing by amiRNA had been established in fertile transgenic soybean. We demonstrated that knockdown of storage protein shifted the distribution of nitrogen sources in soybean seeds.

Abstract

Artificial microRNAs (amiRNAs) were designed using the precursor sequence of the endogenous soybean (Glycine max L. Merrill) miRNA gma-miR159a and expressed in transgenic soybean plants to suppress the biosynthesis of 7S globulin, which is one of the major storage proteins. Seed-specific expression of these amiRNAs (amiR-7S) resulted in a strong suppression of 7S globulin subunit genes and decreased accumulation of the 7S globulin subunits in seeds. Thus, the results demonstrate that a platform for gene silencing by amiRNA was first developed in fertile transgenic soybean plants. There was no difference in nitrogen, carbon, and lipid contents between amiR-7S and control seeds. Four protein fractions were collected from defatted mature seeds on the basis of solubility at different pH to examine the distribution of nitrogen sources and compensatory effects. In the whey and lipophilic fractions, nitrogen content was similar in amiR-7S and control seeds. Nitrogen content was significantly decreased in the major soluble protein fraction and increased in the residual fraction (okara) of the amiR-7S seeds. Amino acid analysis revealed that increased nitrogen compounds in okara were proteins or peptides rather than free amino acids. Our study indicates that the decrease in 7S globulin subunits shifts the distribution of nitrogen sources to okara in transgenic soybean seeds.

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Acknowledgments

We thank Y. Kitsui and M. Suzuki (Hokkaido University), and E. Okuda (Kyoto University) for technical assistance. This work was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan (23580002) and by CREST of the Japan Science and Technology Agency (JAT).

Conflict of interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Graduate School of Agriculture, Hokkaido University, Kita9 Nishi9, Kita-ku, Sapporo, Hokkaido, 060-8589, Japan

    Tetsuya Yamada, Yoshihiro Mori, Keisuke Kitamura & Jun Abe

  2. School of Agriculture, Hokkaido University, Kita9 Nishi9, Kita-ku, Sapporo, Hokkaido, 060-8589, Japan

    Kazuho Yasue

  3. Graduate School of Agriculture, Kyoto University, Uji, Kyoto, 611-011, Japan

    Nobuyuki Maruyama

Authors
  1. Tetsuya Yamada
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  2. Yoshihiro Mori
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  3. Kazuho Yasue
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  4. Nobuyuki Maruyama
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  5. Keisuke Kitamura
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  6. Jun Abe
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Corresponding author

Correspondence to Tetsuya Yamada.

Additional information

Communicated by Kinya Toriyama.

T. Yamada and Y. Mori contributed equally to this work.

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Yamada, T., Mori, Y., Yasue, K. et al. Knockdown of the 7S globulin subunits shifts distribution of nitrogen sources to the residual protein fraction in transgenic soybean seeds. Plant Cell Rep 33, 1963–1976 (2014). https://doi.org/10.1007/s00299-014-1671-y

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  • DOI: https://doi.org/10.1007/s00299-014-1671-y

Keywords

  • Glycine max
  • AmiRNA
  • Gene silencing
  • Storage protein
  • Protein content
  • Amino acids