Cold stress in Lotus japonicus: a transcriptomic and physiological approach

CALZADILLA P.I.; ESCARAY, F.J; SIGNORELLI S; MAIALE SANTIAGO; MENÉNDEZ A.; MONZA J; RUIZ O.A.

Taller; VI Taller de Lotus spp; 2016

Members of the Lotus genus are important as agricultural forage sources under marginal environmental conditions given their high nutritional value and tolerance to various abiotic stresses. However, the dry matter production and regrowth rate of cultivable Lotus spp. is drastically reduced during colder seasons, while their response to such conditions is not well studied. High-throughput RNA sequencing was used to identify and classify 1.077 differentially expressed genes, after 24 h of cold stress in L. japonicus. Of them, 713 were up-regulated and 364 were down-regulated. Up-regulated genes were principally related to lipid, cell wall, phenylpropanoid, sugar, and proline regulation, while down-regulated genes affected the photosynthetic process and chloroplast development. Together, a total of 41 cold-inducible transcription factors were identified, including members of the AP2/ERF, NAC, MYB, and WRKY families; two of them were described as putative novel transcription factors. Altogether, this is the first transcriptome profiling of the model legume L. japonicus under cold stress. Results obtained were used to identify key components in the cold acclimation response of two L. japonicus ecotypes, with contrasting response to low temperature (MG-1 and MG-20). Oxidative and nitrosative stress indicators, proline content and photosynthesis parameters were evaluated on 3-weeks-old plants of these ecotypes at different times (first, fourth and seventh day) of stress (5-9 ± 1 °C) and control (21-25 ± 1 °C) treatments. Results showed that a chilling-induced redox imbalance was evidenced during the cold stress treatment and, that a stress-induced photoinhibition differentially influenced both ecotypes, being MG-1 more affected than MG-20. Interestingly, measurements of the protein levels of the D1 and D2 PSII reaction centre proteins, suggested that D2 protein may be involved in the acclimation response of L. japonicus to low temperature. Our results may provide a deeper insight into the chilling tolerance mechanisms of the Lotus genus, and provide candidate genes and putative metabolic pathways for breeding modified species of forage legumes that more readily acclimate to low temperatures.