DgbZIP3 interacts with DgbZIP2 to increase the expression of<i>DgPOD</i>for cold stress tolerance in chrysanthemum

Abstract

The bZIP transcription factor plays a very important role in abiotic stresses, e.g. drought, salt, and low-temperature stress, but the mechanism of action at low temperature is still unclear. In this study, overexpression of <i>DgbZIP3</i> led to increased tolerance of chrysanthemum (<i>Chrysanthemum morifolium</i> Ramat.) to cold stress, whereas antisense suppression of <i>DgbZIP3</i> resulted in decreased tolerance. Electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation (ChIP), luciferase complementary imaging analysis (LCI), and dual-luciferase reporter gene detection (DLA) experiments indicated that DgbZIP3 directly bound to the promoter of <i>DgPOD</i> and activated its expression. DgbZIP2 was identified as a DgbZIP3-interacting protein using yeast two-hybrid, co-immunoprecipitation, LCI, and bimolecular fluorescence complementation assays. Overexpression of <i>DgbZIP2</i> led to increased tolerance of chrysanthemum to cold stress, whereas antisense suppression of <i>DgbZIP2</i> resulted in decreased tolerance. A ChIP-qPCR experiment showed that DgbZIP2 was highly enriched in the promoter of <i>DgPOD</i>, while DLA, EMSA, and LCI experiments further showed that DgbZIP2 could not directly regulate the expression of <i>DgPOD</i>. The above results show that DgbZIP3 interacts with DgbZIP2 to regulate the expression of <i>DgPOD</i> to promote an increase in peroxidase activity, thereby regulating the balance of reactive oxygen species and improving the tolerance of chrysanthemum to low-temperature stress.