Genome-wide identification of CML gene family in Salix matsudana and functional verification of SmCML56 in tolerance to salts tress

Calmodulin-like protein (CML) mediates Ca2+ signaling in response to abiotic stress. It has been shown that manipulating this signaling can improve crop stress resistance. However, the CML family in Willow has not been comprehensively and deeply studied. In this study, 157 SmCML genes were identified on the whole genome of Salix matsudana using bioinformatics method. Phylogenetic analysis showed that CML homologs between S. matsudana and Arabidopsis thaliana shared close relationships. The identified SmCML genes were distributed on 41 chromosomes. Analysis of cis-acting elements indicated that SmCMLs play an important role in plant hormone signal transduction and environmental stress response. SmCML56 gene was successfully cloned from S. matsudana and overexpressed in A. thaliana was constructed by flower dip method, and overexpressed in willow was constructed by Agrobacterium rhizogenes K599 mediated genetic transformation of willow hairy roots. Phenotypic, physiological and biochemical analysis confirmed that overexpression of SmCML56 significantly increased the tolerance of plants to salt. At the same time, VIGS experiment showed that the tolerance of silenced plants to salt stress decreased. The results of this study increased the understanding and characterization of SmCML genes in willow and will be a rich resource for further studies to investigate SmCML protein function in various developmental processes of willow. It provided a reference for related calmodulin-like studies in other perennial species.