Arabidopsis thaliana ASN2 encoding asparagine synthetase is involved in the control of nitrogen assimilation and export during vegetative growth

• Published in: Plant, cell and environment cell and environment, 2013-02, Vol.36 (2), p.328-342
• Main Authors: GAUFICHON, LAURE, MASCLAUX‐DAUBRESSE, CÉLINE, TCHERKEZ, GUILLAUME, REISDORF‐CREN, MICHÈLE, SAKAKIBARA, YUKIKO, HASE, TOSHIHARU, CLÉMENT, GILLES, AVICE, JEAN‐CHRISTOPHE, GRANDJEAN, OLIVIER, MARMAGNE, ANNE, BOUTET‐MERCEY, STÉPHANIE, AZZOPARDI, MARIANNE, SOULAY, FABIENNE, SUZUKI, AKIRA
• Format: Article
• Language: English
• Subjects: Agricultural sciences; amino acid synthesis and metabolism; Arabidopsis - enzymology; Arabidopsis - genetics; Arabidopsis - growth & development; Arabidopsis Proteins - genetics; Arabidopsis Proteins - metabolism; Aspartate-Ammonia Ligase - genetics; Aspartate-Ammonia Ligase - metabolism; Biological and medical sciences; Biological Transport; cellular localization; DNA, Bacterial - genetics; Fundamental and applied biological sciences. Psychology; Gases - metabolism; Gene Expression Profiling; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Plant; Genes, Plant - genetics; Life Sciences; Metabolome; Mutagenesis, Insertional - genetics; Mutation - genetics; Nitrogen - metabolism; nitrogen assimilation and translocation; Phenotype; Phloem - enzymology; Photosynthesis; Plant Leaves - metabolism; reverse genetics; RNA, Messenger - genetics; RNA, Messenger - metabolism
• ISSN: 0140-7791; 1365-3040
• DOI: 10.1111/j.1365-3040.2012.02576.x

ABSTRACT We investigated the function of ASN2, one of the three genes encoding asparagine synthetase (EC 6.3.5.4), which is the most highly expressed in vegetative leaves of Arabidopsis thaliana. Expression of ASN2 and parallel higher asparagine content in darkness suggest that leaf metabolism involves ASN2 for asparagine synthesis. In asn2‐1 knockout and asn2‐2 knockdown lines, ASN2 disruption caused a defective growth phenotype and ammonium accumulation. The asn2 mutant leaves displayed a depleted asparagine and an accumulation of alanine, GABA, pyruvate and fumarate, indicating an alanine formation from pyruvate through the GABA shunt to consume excess ammonium in the absence of asparagine synthesis. By contrast, asparagine did not contribute to photorespiratory nitrogen recycle as photosynthetic net CO2 assimilation was not significantly different between lines under both 21 and 2% O2. ASN2 was found in phloem companion cells by in situ hybridization and immunolocalization. Moreover, lack of asparagine in asn2 phloem sap and lowered 15N flux to sinks, accompanied by the delayed yellowing (senescence) of asn2 leaves, in the absence of asparagine support a specific role of asparagine in phloem loading and nitrogen reallocation. We conclude that ASN2 is essential for nitrogen assimilation, distribution and remobilization (via the phloem) within the plant. This work demonstrates the specific functions of ASN2 encoding asparagine synthetase in Arabidopsis thaliana at a vegetative phase. ASN2 mediates assimilation of ammonium into asparagine within the phloem companion cells for use to nitrogen translocation in Arabidopsis. The lack of asparagine due to the ASN2 disruption causes an alteration of nitrogen distribution into amino acids in both leaves and phloem. The delayed senescence of the ASN2‐deficient Arabidopsis lines provides evidence for the ASN2‐mediated synthesis of asparagine that serves as a specific amide for the nitrogen remobilization.