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Arabidopsis mutants lacking asparaginases develop normally but exhibit enhanced root inhibition by exogenous asparagine

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Abstract

Asparaginase catalyzes the degradation of l-asparagine to l-aspartic acid and ammonia, and is implicated in the catabolism of transported asparagine in sink tissues of higher plants. The Arabidopsis genome includes two genes, ASPGA1 and ASPGB1, belonging to distinct asparaginase subfamilies. Conditions of severe nitrogen limitation resulted in a slight decrease in seed size in wild-type Arabidopsis. However, this response was not observed in a homozygous T-DNA insertion mutant where ASPG genes had been inactivated. Under nitrogen-sufficient conditions, the ASPG mutant had elevated levels of free asparagine in mature seed. This phenotype was observed exclusively under conditions of low illumination, when a low ratio of carbon to nitrogen was translocated to the seed. Mutants deficient in one or both asparaginases were more sensitive than wild-type to inhibition of primary root elongation and root hair emergence by l-asparagine as a single nitrogen source. This enhanced inhibition was associated with increased accumulation of asparagine in the root of the double aspga1-1/-b1-1 mutant. This indicates that inhibition of root growth is likely elicited by asparagine itself or an asparagine-derived metabolite, other than the products of asparaginase, aspartic acid or ammonia. During germination, a fusion between the ASPGA1 promoter and beta-glucuronidase was expressed in endosperm cells starting at the micropylar end. Expression was initially high throughout the root and hypocotyl, but became restricted to the root tip after three days, which may indicate a transition to nitrogen-heterotrophic growth.

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Acknowledgments

We thank Douglas Johnson at the University of Ottawa for providing the pCAMBIA1302tZ-PrxN-GUS vector. We are grateful to Larry Stitt at the Department of Epidemiology and Biostatistics, University of Western Ontario for advice on statistical analysis, and Rey Interior, at the Advanced Protein Technology Centre, Hospital for Sick Children for amino acid analyses. We are indebted to staff at the Southern Crop Protection and Food Research Centre, Ida van Grinsven for DNA sequencing, Tim McDowell for elemental analysis and Alex Molnar for preparation of figures. Funding was provided by the Discovery Program of the Natural Sciences and Engineering Research Council.

Conflict of interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Department of Biology, University of Western Ontario, London, ON, Canada

    Ana Ivanov & Frédéric Marsolais

  2. Agriculture and Agri-Food Canada, Southern Crop Protection and Food Research Centre, 1391 Sandford Street, London, ON, N5V 4T3, Canada

    Ana Ivanov, Alexander Kameka, Agnieszka Pajak, Luanne Bruneau, Cinta Hernández-Sebastià & Frédéric Marsolais

  3. Agriculture and Agri-Food Canada, Southern Crop Protection and Food Research Centre, Research Farm, Delhi, ON, Canada

    Ronald Beyaert

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  1. Ana Ivanov
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  2. Alexander Kameka
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  3. Agnieszka Pajak
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Correspondence to Frédéric Marsolais.

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726_2011_973_MOESM1_ESM.pdf

Supplementary material 1 (PDF 55 kb) Table 1 C, S and N content of mature seed of wild-type and aspga1-1/-b1-1 grown under normal (115 μmol photons m−2 sec−1) or low (70 μmol photons m−2 sec−1) illumination. Table 2 Total amino acid profile of mature seeds of wild-type and aspga1-1/-b1-1 grown under low light conditions

726_2011_973_MOESM2_ESM.pdf

Supplementary material 2 (PDF 35 kb) Figure 1 Graphs of D × G interactions from ANOVA of values in Tables 2 (a) and 3 (b)

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Ivanov, A., Kameka, A., Pajak, A. et al. Arabidopsis mutants lacking asparaginases develop normally but exhibit enhanced root inhibition by exogenous asparagine. Amino Acids 42, 2307–2318 (2012). https://doi.org/10.1007/s00726-011-0973-4

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  • DOI: https://doi.org/10.1007/s00726-011-0973-4

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