Genome-Wide Two-Component System Genes of Cytokinin Analysis and Their Functions of Drought Stress Response in Pinus tabuliformis

Objective: Cytokinin signaling plays an important role in plant growth and development, but cytokinin two-component signaling system (TCS) genes have not been comprehensively reported in conifers. We aimed to identify and analyze the TCS family in Pinus tabuliformis and explore their roles in drought response. Methods: PtTCS genes were identified by BLASTP using Arabidopsis TCS sequences and validated by conserved domain analysis. Phylogenetic, structural, cis-element, GO enrichment, and expression profiling under drought were performed, with qRT-PCR validation. Results: Twenty-two PtTCS genes were identified (three CHKs, four HPs, eight type-A RRs, seven type-B RRs). Most proteins retained canonical domains, though some key residue variations were noted. Five genes (PtCHK2, PtHP1, PtHP4, PtRR2, PtRR4) were significantly upregulated under drought, opposite to the pattern in angiosperms, and rewatering restored expression. Conclusions: The drought-induced upregulation in P. tabuliformis contrasts with angiosperm downregulation, suggesting a distinct cytokinin-mediated drought response in conifers. This study provides the first comprehensive TCS inventory in a conifer and a foundation for functional studies in gymnosperms.