Genome-wide analysis of 14-3-3 genes in sugarcane and functional analysis of SsGF2.1 in response to chilling stress

The 14-3-3 proteins are a family of highly conserved acidic proteins that play a vital role in regulating stress response. Saccharum spontaneum is a founding species that contributes stress tolerance to the genetic background of modern sugarcane (Saccharum spp. Hybrids), an important crop for sugar and biofuel production. Members of the 14-3-3 family in S. spontaneum, especially those involved in response to chilling stress, have not yet been analyzed. In this study, a total of 24 SsGF genes with characteristic 14-3-3 domains were identified in the genome of S. spontaneum AP85-441. These SsGF genes are unevenly distributed on 11 chromosomes and could be divided into 2 types (Epsilon and non -Epsilon types). Colinear analysis indicated that the expansion of the SsGF genes in S. spontaneum is mainly due to gene fragment duplication events. Most SsGF genes contain abiotic stress response elements in their promoter regions. RNA-seq analysis demonstrated that SsGF2.1-2.4 were highly expressed in growth and development, circadian rhythms and cold responsiveness, suggesting that these 4 SsGF2s may be key 14-3-3 genes. Furthermore, putative SsGF2.1-interacting proteins were detected by screening a yeast two -hybrid cDNA library. Three of them, SsMYBS3, SsCBSX2-1, and SsCBSX2-2, were verified for their interactions with SsGF2.1 and found to have similar expression patterns with SsGF2.1. These findings provide important candidate genes for genetically enhancing cold -tolerance in sugarcane and lay a foundation for understanding the molecular mechanism of sugarcane response to chilling stress.