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Barley (Hordeum vulgare L.) inositol monophosphatase: gene structure and enzyme characteristics

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Abstract

The cellular myo-inositol (Ins) pool is important to many metabolic and signaling pathways in plants. Ins monophosphatase (IMPase; EC 3.1.3.25) activity is essential for the de novo synthesis of myo-Inositol (Ins), and for recycling of Ins in Ins(1,4,5)P3. However, proteins encoded by at least one family of IMP genes also have l-galactose-1-P phosphatase activity important to ascorbic acid synthesis, indicating a bifunctionality that links these two branches of carbon metabolism. As part of research into the regulation of Ins synthesis and supply during seed development, the barley IMP-1 gene and gene products were studied. The 1.4 kb barley IMP-1 promoter contains one low temperature response element (RE), two heat shock REs, one gibberellin and two auxin REs, and five sugar REs. Barley IMP-1 is expressed in all tissues assayed, and expression levels were not greatly altered by abiotic stress treatments. Reduced use of Ins for Ins P6 synthesis in developing seed of barley low phytic acid (lpa) mutants results in Ins accumulation, and IMP-1 expression is reduced in proportion to the increase in Ins level. The barley recombinant enzyme had a lower Km, indicating higher affinity, for d/l-Ins(3)P1 (Km = 9.7 μM) as compared with reported Km (Ins P1) values for other eukaryotic IMPases (43–330 μM) or with a reported Km (l-Gal-1P) of 150 μM for a kiwifruit (Actinidia deliciosa) enzyme. These and other data indicate that the barley IMP-1 gene is regulated at least in part in response to Ins metabolic needs, and that the enzyme it encodes displays catalytic properties well suited for a role in Ins synthesis, in addition to other roles as an l-gal-1-P phosphatase important to ascorbate synthesis, or as an IMPase important to Ins(1,4,5)P3 signal recycling.

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Notes

  1. With the exception of the use of “d/l” mixtures used in enzyme assays, all Ins phosphates will be referred to here using the “d”-numbering convention; d-Ins(3)P1 is identical to l-Ins(1)P1.

Abbreviations

CTAB:

Hexadecyltrimethylammonium bromide

HEPES:

N-2-Hydroxyethylpiperazine-N′-2-ethane-sulfonic acid

Ins:

myo-Inositol

lpa :

Low phytic acid

Ins P6 :

myo-Inositol-1,2,3,4,5,6-hexakisphosphate

IPTG:

Isopropyl β-d-thiogalactoside

MIPS:

myo-Inositol-3-phosphate synthase

IMPase:

myo-Inositol monophosphatase

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Acknowledgements

The authors would like to thank Justin Wheeler and David Bowen for help with the greenhouse work, and Phillip Bregitzer and Ann Sturbaum for help with the BAC library filter hybridization. This work was supported in part by the National Science Foundation (Project No. 0110748).

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Author notes

  1. Jianming Fu

    Present address: Department of Agronomy, Kansas State University, Manhattan, KS, 66506, USA

  2. Kevin Peterson

    Present address: Mayo Clinic, 1216 Second Street SW, Rochester, MN, 55902, USA

  3. Mary Guttieri & Edward Souza

    Present address: USDA-ARS, 1680 Madison Ave., Wooster, OH, 44691, USA

Authors and Affiliations

  1. Research and Extension Center, University of Idaho, 1699 S 2700 W, Aberdeen, ID, 83210, USA

    Jianming Fu, Mary Guttieri & Edward Souza

  2. USDA-ARS, 1691 South 2700 West, Aberdeen, ID, 83210, USA

    Kevin Peterson & Victor Raboy

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  1. Jianming Fu
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  2. Kevin Peterson
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  5. Victor Raboy
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Corresponding author

Correspondence to Victor Raboy.

Additional information

The barley IMP-1 cDNA and protein sequence accession number is AY460570. Accession numbers for the barley genomic sequence including promoter and coding region are DQ145527 and DQ145528. These sequences will be available in GenBank upon the publication of this paper.

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Fu, J., Peterson, K., Guttieri, M. et al. Barley (Hordeum vulgare L.) inositol monophosphatase: gene structure and enzyme characteristics. Plant Mol Biol 67, 629–642 (2008). https://doi.org/10.1007/s11103-008-9343-3

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  • DOI: https://doi.org/10.1007/s11103-008-9343-3

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