Gene Expression and Interaction Analysis of FsWRKY4 and FsMAPK3 in Forsythia suspensa

Xinjie Tan; Jiaxi Chen; Jiaqi Zhang; Guangyang Guo; Hongxiao Zhang; Xingli Zhao; Shufang Lv; Huawei Xu; Dianyun Hou

doi:10.3390/plants12193415

Gene Expression and Interaction Analysis of FsWRKY4 and FsMAPK3 in Forsythia suspensa

Plants (Basel). 2023 Sep 28;12(19):3415. doi: 10.3390/plants12193415.

Authors

Xinjie Tan¹, Jiaxi Chen¹, Jiaqi Zhang¹, Guangyang Guo¹, Hongxiao Zhang¹, Xingli Zhao¹, Shufang Lv¹, Huawei Xu¹, Dianyun Hou¹

Affiliation

¹ College of Agriculture, Henan University of Science and Technology, Luoyang 471032, China.

Abstract

Forsythia suspensa is a deciduous shrub that belongs to the family Myrtaceae, and its dried fruits are used as medicine. F. suspensa contains several secondary metabolites, which exert pharmacological effects. One of the main active components is forsythin, which exhibits free radical scavenging, antioxidant, anti-inflammatory, and anti-cancer effects. Mitogen-activated protein kinase (MAPKs) can increase the activity of WRKY family transcription factors in a phosphorylated manner, thereby increasing the content of secondary metabolites. However, the mechanism of interaction between MAPKs and WRKYs in F. suspensa remains unclear. In this study, we cloned the genes of FsWRKY4 and FsMAPK3, and performed a bioinformatics analysis. The expression patterns of FsWRKY4 and FsMAPK3 were analyzed in the different developmental stages of leaf and fruit from F. suspensa using real-time fluorescence quantitative PCR (qRT-PCR). Subcellular localization analysis of FsWRKY4 and FsMAPK3 proteins was performed using a laser scanning confocal microscope. The existence of interactions between FsWRKY4 and FsMPAK3 in vitro was verified by yeast two-hybridization. Results showed that the cDNA of FsWRKY4 (GenBank number: OR566682) and FsMAPK3 (GenBank number: OR566683) were 1587 and 522 bp, respectively. The expression of FsWRKY4 was higher in the leaves than in fruits, and the expression of FsMAPK3 was higher in fruits but lower in leaves. The subcellular localization results indicated that FsWRKY4 was localized in the nucleus and FsMAPK3 in the cytoplasm and nucleus. The prey vector pGADT7-FsWRKY4 and bait vector pGBKT7-FsMAPK3 were constructed and co-transferred into Y2H Glod yeast receptor cells. The results indicated that FsWRKY4 and FsMAPK3 proteins interact with each other in vitro. The preliminary study may provide a basis for more precise elucidation of the synthesis of secondary metabolites in F. suspensa.

Keywords: Forsythia suspensa; FsMAPK3; FsWRKY4; gene expression; interaction.

Grants and funding

This research was supported by the National Natural Science Foundation of China (U1404829); Key project at central government level: The ability establishment of sustainable use for valuable Chinese medicine resources (2060302); and the Natural Science Foundation of Henan Province (202300410151). This research was also supported by a special fund for the construction of technical systems of the traditional Chinese medicine industry in Henan Province, from the Henan Province Chinese medicine industry science and technology commissioner service group.