关键词: 山新杨, 舞毒蛾, 茉莉酸, 酵母双杂交, 植物防御

Abstract:

Objective: This study aims to investigate the expression characteristics and biological functions of the PdbpetA gene in Populus davidiana × P. bolleana, and elucidate the molecular mechanism by which this gene is activated and regulates plant defense responses upon induction by the Lymantria dispar oral effector Armet. Method: The open reading frame (ORF) sequence of the PdbpetA gene from P. davidiana × P. bolleana was obtained by PCR cloning. Using techniques such as restriction enzyme digestion and homologous recombination, the PdbpetA gene sequence was constructed into plant transient expression, subcellular localization, and bimolecular fluorescence complementation (BiFC) vectors. The transient transformation system was used to analyze the subcellular localization of PdbpetA protein in tobacco cells. Transient expression of the PdbpetA gene in transgenic P. davidiana × P. bolleana was achieved through Agrobacterium-mediated transient transformation. The quantitative real-time RT-PCR was employed to examine the tissue-specific expression patterns of the PdbpetA gene in P. davidiana × P. bolleana. The relative expression levels of jasmonic acid biosynthesis-related genes PdbJAR1, PdbAOC, and PdbLOX2 were compared between control and transgenic plants. Additionally, proteins interacting with PdbpetA were screened using a yeast two-hybrid (Y2H) system. Result: The ORF of the PdbpetA gene is 696 bp in length and encodes 231 amino acids. The gene exhibited the highest expression level in mature leaves and the protein was localized to both the nucleus and cell membrane. Transient overexpression of PdbpetA in P. davidiana × P. bolleana reached its peak at 48 hours after transformation, whereas transient suppression led to significantly reduced expression at 36 hours. After L. dispar larvae feeding on PdbpetA-overexpressing plants, the relative expression levels of jasmonic acid biosynthesis-related genes PdbJAR1, PdbAOC, and PdbLOX2 were significantly up-regulated. However, these JA synthesis genes were markedly down-regulated in plants with transiently suppressed PdbpetA. Protein interaction analysis revealed that PdbpetA interacted with PdbKunitz and PdbrDNA. Conclusion: The PdbpetA gene of P. davidiana × P. bolleana not only participates in plant jasmonic acid synthesis, but also likely participates in plant defense responses through interactions with PdbKunitz and PdbrDNA.

Key words: Populus davidiana × P. bolleana, Lymantria dispar, jasmonic acid, yeast two-hybrid system, plant defense