Disruption of Asparagine Synthetase Is Associated to Increased Biomass in <i>Lotus japonicus</i>

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.