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Hordein promoter methylation and transcriptional activity in wild-type and mutant barley endosperm

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Abstract

B- and C-hordein gene transcription is severely reduced in the endosperm of the regulatory barley mutantlys3a, and this is correlated with persistent hypermethylation of the promoters. In contrast, D-hordein is expressed at normal levels in the mutant. To confirm the connection between methylation and transcriptional activity, a genomic D-hordein clone was isolated and sequenced. The nucleotide composition of the promoter region revealed a CpG island and methylation analysis, using bisulphite treatment of genomic DNA, confirmed that the D-hordein promoter is unmethylated in endosperm and leaf tissue. Immunocytochemical studies localized D-hordein to the reticular component of protein bodies in both the wild-type Bomi andlys3a. Transient expression ofGUS reporter gene constructs in barley endosperm, following transfection by particle bombardment revealed the D-hordein promoter to be 3–5 fold more active than B-or C-hordein promoters. Comparison of transient expression in Bomi andlys3a endosperm demonstrated that the activities of the unmethylated D-hordein and theHor1-14 C-hordein promoters were equivalent, while the activities in the mutant of theHor1-17 C-hordein and theHor2-4 B-hordein promoters were reduced two- and tenfold, respectively. Methylation of plasmids in vitro prior to expression severely inhibited B- and D-hordein promoter activities. Based on these observations two categories of promoters for endosperm-specific expression of storage proteins are recognized and a model involving methylation and modulation of chromatin structure in the regulation by theLys3 gene is presented.

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Author notes

  1. Martin Müller

    Present address: Max-Planck-Institut für Züchtungsforschung, Carl-von-Linné-Weg 10, D-50829, Köln, Germany

Authors and Affiliations

  1. Department of Physiology, Carlsberg Laboratory, Gamle Carlsbergvej 10, DK-2500, Copenhagen Valby, Denmark

    Mikael Blom Sørensen, Martin Müller & David Simpson

  2. Department of Crop and Soil Sciences, Washington State University, 99164, Pullman, WA, USA

    Mikael Blom Sørensen

  3. CSIRO Division of Plant Industry, GPO Box 1600, 2601, Canberra, ACT, Australia

    John Skerritt

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  1. Mikael Blom Sørensen
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  2. Martin Müller
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  4. David Simpson
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Communicated by H. Saedler

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Sørensen, M.B., Müller, M., Skerritt, J. et al. Hordein promoter methylation and transcriptional activity in wild-type and mutant barley endosperm. Molec. Gen. Genet. 250, 750–760 (1996). https://doi.org/10.1007/BF02172987

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  • DOI: https://doi.org/10.1007/BF02172987

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