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Functions of the CCCH type zinc finger protein OsGZF1 in regulation of the seed storage protein GluB-1 from rice

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Abstract

Glutelins are the most abundant storage proteins in rice grain and can make up to 80 % of total protein content. The promoter region of GluB-1, one of the glutelin genes in rice, has been intensively used as a model to understand regulation of seed-storage protein accumulation. In this study, we describe a zinc finger gene of the Cys3His1 (CCCH or C3H) class, named OsGZF1, which was identified in a yeast one-hybrid screening using the core promoter region of GluB-1 as bait and cDNA expression libraries prepared from developing rice panicles and grains as prey. The OsGZF1 protein binds specifically to the bait sequence in yeast and this interaction was confirmed in vitro. OsGZF1 is predominantly expressed in a confined domain surrounding the scutellum of the developing embryo and is localised in the nucleus. Transient expression experiments demonstrated that OsGZF1 can down-regulate a GluB-1-GUS (β-glucuronidase) reporter and OsGZF1 was also able to significantly reduce activation conferred by RISBZ1 which is a known strong GluB-1 activator. Furthermore, down-regulation of OsGZF1 by an RNAi approach increased grain nitrogen concentration. We propose that OsGZF1 has a function in regulating the GluB-1 promoter and controls accumulation of glutelins during grain development.

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Acknowledgments

This research was supported by the Programme for Strategic Scientific Alliances (04-PSA-BD-04) between China and the Netherlands of the Royal Netherlands Academy of Arts and Sciences (KNAW) for MW, PBFO, ZZ and the KNAW China Exchange Programmes (04CDP022, 06CDO033, 07CDP005, 08CDP042) for PBFO, MW, ZZ and YC.

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  1. Yi Chen

    Present address: Rothamsted Research, West Common, Harpenden, AL5 2JQ, UK

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  1. Sylvius Laboratory, Institute of Biology (IBL), Leiden University, Sylviusweg 72, 2333 BE, PO Box 9505, 2300 RA, Leiden, The Netherlands

    Yi Chen, Aijun Sun, Mei Wang & Pieter B. F. Ouwerkerk

  2. State Key Laboratory of Plant Genomics, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, No.1 West Beichen Road, Chaoyang District, Beijing, 100101, China

    Aijun Sun & Zhen Zhu

  3. SU BioMedicine TNO, Utrechtseweg 48, 3704 HE Zeist, PO Box 360, 3700 AJ, Zeist, The Netherlands

    Mei Wang

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11103_2013_158_MOESM1_ESM.tif

Fig. S1 cDNA and protein sequence of OsGZF1. The zinc finger motifs are underlined and the Cys and His residues within the zinc finger motifs are marked with squares. (TIFF 1731 kb)

11103_2013_158_MOESM2_ESM.tif

Fig. S2 Yeast activation assay. pAS2-1/OsGZF1 and empty control vector pAS2-1 in yeast strain PJ69-4A and grown under histidine (a) or adenine selection (b). As a result, the Gal4p/OsGZF1 fusion can not activate the HIS3 or ADE2 reporters. Growth was monitored after 7 days at 30 °C. (TIFF 1833 kb)

11103_2013_158_MOESM3_ESM.tif

Fig. S3 Northern blot analysis of OsGZF1/RNAi plants showing no obvious change of the expression level of GluB-1 mRNA in rice grains (10 DAF). (TIFF 885 kb)

Fig. S4 Comparison of OsGZF1 and PEI1. Zinc finger motifs are underlined. (TIFF 1679 kb)

Supplementary material 5 (DOC 74 kb)

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Chen, Y., Sun, A., Wang, M. et al. Functions of the CCCH type zinc finger protein OsGZF1 in regulation of the seed storage protein GluB-1 from rice. Plant Mol Biol 84, 621–634 (2014). https://doi.org/10.1007/s11103-013-0158-5

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