Disruption of Asparagine Synthetase Is Associated to Increased Biomass in Lotus japonicus

Asparagine (Asn) constitutes the major form of nitrogen translocated within Lotus japonicus plants. In this work we use knock-out (KO) LORE1 mutants-deficient in the asparagine synthetase gene (LjASN1), which is the most highly expressed ASN gene in plants grown under non-symbiotic (NS) conditions, but much less expressed under symbiotic (S) conditions. The analysis of two different Ljasn1 homozygous mutant lines grown under NS or S conditions indicated that a much higher biomass was produced in Ljasn1 mutants grown under NS conditions compared to the WT (wild-type), whereas little difference with the WT was observed in mutant plants under S conditions. Metabolomic analysis revealed that Ljasn1 mutant plants are quite distinct to WT plants when grown under NS conditions, but not under S conditions. Asn levels were considerably reduced in Ljasn1 mutant plants compared to the WT when plants were grown under NS but not under S conditions. A general decrease in amino acids and an increase in carbon compounds, such as sugars and oxo-acids, was detected in NS roots and shoots, respectively, which may explain the growth phenotypes observed. RNAseq analysis showed changes related to oxidative metabolism under NS conditions, and C/N metabolism under S conditions. The data indicate that the LjASN1 deficiency produces important changes in the C/N balance and metabolite allocation of L. japonicus plants resulting in higher biomass content and lower Asn levels, two interesting traits for biotechnological crops engineering.