The interaction of CsWRKY4 and CsOCP3 with CsICE1 regulates CsCBF1/3 and mediates stress response in tea plant (Camellia sinensis)

Abstract

Low temperature stress limits the geographical distribution of tea plants (Camellia sinensis) and the yield and quality of tea. CsICE1 is a crucial regulator of tea plant’s cold signaling pathway; however, the regulatory mechanism of CsICE1 is still unclear. In this study, we found that the N-terminus of CsICE1 contains a transcriptional activation domain and a region of protein interaction. Yeast two-hybrid and bimolecular fluorescence complementation experiments confirmed that CsWRKY4 and CsOCP3 interacted with CsICE1 in the nucleus. Like CsICE1, CsWRKY4 and CsOCP3 were localized in the nucleus, and CsOCP3 was also targeted to the chloroplast. The expression of CsWRKY4 and CsOCP3 was downregulated under cold stress, unlike that of CsICE1. Under biotic stresses and salicylic acid treatment, the expression of all three genes was increased, whereas jasmonic acid treatment resulted in decreased expression of all three genes. Dual-luciferase transient assays revealed that CsICE1 promoted the expression of CsCBF1 and CsCBF3, whereas CsWRKY4 and CsOCP3 inhibited the expression of CsCBF1 and CsCBF3 and attenuated their induction by CsICE1. Taken together, these results suggest that CsWRKY4 and CsOCP3 interact with CsICE1 and regulate CsCBF1/3, thus mediating the stress response in tea plant. Furthermore, these results highlight that CsICE1 is not only a key component in the low temperature signal response pathway, but it may also serve as a point of confluence for cold and other signaling pathways.