Transcriptional Profiling of BpWRKY49 Reveals Its Role as a Master Regulator in Stress Signaling Pathways in Birch (Betula platyphylla)

Abstract

The WRKY family of transcription factors (TFs) is one of the most diverse families in plants, playing crucial roles in various plant growth and stress response processes. Asian white birch (Betula platyphylla) is a globally distributed tree species that holds ecological, medical, and economic significance. However, the regulatory mechanisms of WRKY TFs in birch remain poorly understood. Herein, we cloned and characterized the BpWRKY49 gene from birch. Through bioinformatics analyses, we revealed the potential involvement of BpWRKY49 in both biotic and abiotic stress responses. In addition, BpWRKY49 was found to be localized in the nucleus and exhibited transcriptional activity in yeast. Transactivation assays further confirmed that BpWRKY49 exhibited transcriptional activity at its C-terminal end. Notably, our binding specificity assays demonstrated the specific interaction of BpWRKY49 with the W-box cis element in vitro. Furthermore, tissue-specific expression analysis demonstrated that BpWRKY49 exhibited the highest expression level in the roots. Real-time quantitative PCR (RT-qPCR) analysis of birch plants subjected to salt and drought treatments revealed that BpWRKY49 displayed significant 30-fold and 10-fold upregulations under salt and drought stress conditions, respectively. DAP-seq analysis of BpWRKY49 identified a total of 21,832 peaks, with 3477 occurring in the promoter region of genes. Gene ontology (GO) enrichment analysis highlighted prominent terms related to defense against biotic stress, followed by terms associated with abiotic stress and development. Y1H assays of three genes provided evidence for the binding ability of BpWRKY49 to the promoters of BpPUB21, BpBTL15, and BpHIP47 in vitro. Collectively, our findings strongly suggest that BpWRKY49 possesses diverse functions and may activate multiple genes to contribute to various biological processes, including salt stress tolerance, in birch.