Identification and Expression Analysis of CDPK Family inEriobotrya japonica, revealsEjCDPK25in Response to Freezing Stress in Fruitlets

Abstract The fruitlets of loquat ( Eriobotrya japonica Lindl.) are susceptible to freezing injury due to their developmental cycle encountering winter. Freezing stress severely damages the fruitlets, resulting in loss of fruit yield and quality. Studies have shown that Ca 2+ , as a second messenger, is involved in signal transduction in loquat fruitlets under freezing stress. However, the mechanism of downstream calcium signal transduction in loquat fruitlets under freezing stress is currently unclear. Calcium-depend protein kinase (CDPK) as the most particular calcium sensor family in plants, play an important role in multiple stress signal transduction including freezing. In this study, we identified the loquat CDPK family on a genome-wide scale. A total of 34 EjCDPK genes were identified and studied for basic structural and phylogenetic features. EjCDPKs can be divided into four subgroups phylogenetically. The patterns of exon-intron and protein motif are highly conserved among the subgroups. Collinearity analysis identified several segmental duplicate events in EjCDPK family. RNA-seq based transcription analysis indicated that partial of EjCDPK s differently expressed in response to freezing stress with tissue-specific. Moreover, we preformed correlation analysis between expression value and trait data of loquat fruitlet under freezing stress by weighted co-expression gene network. After that, EjCDPK25 was selected as the candidate because of its potential freezing stress response function. Protein kinase related GO terms were enriched in EjCDPK25 co-expression genes, and then QPCR was performed to examine the target gene’s expression pattern. In addition, EjCDPK25 was cloned to construct overexpression vector to obtain transgenic Arabidopsis plants. Transgenic and wild-type Arabidopsis were suffered freezing stress treatments (-5°C). The results showed that the survival rate of EjCDPK25 overexpressing transgenic Arabidopsis was significantly higher than WT. In summary, this study identified loquat CDPK family firstly, and our data provide significant insights into the evolution and function of loquat CDPKs. Above all, a freezing stress response gene EjCDPK25 was verified can increase the resistance of freezing stress in Arabidopsis .