Genome-wide analysis of the NAC gene family in Qu-1 doubled haploid cell line from Populus simonii × P. nigra and expression patterns under abscisic acid (ABA) treatment.

backgroundPopulus simonii × P. nigra is an important hybrid poplar with strong environmental adaptability, but the molecular basis of its abscisic acid (ABA)-related responses remains unclear. High genomic heterozygosity in poplar also complicates gene family identification and regulatory analysis. In this study, we used the Qu-1 doubled haploid cell line of P. simonii × P. nigra, a genetically homozygous poplar system, to characterize the NAC transcription factor (TF) family and investigate its response to ABA treatment.

resultsA total of 129 Qu-1-PsnNAC genes were identified and classified into 10 subgroups. Phylogenetic, structural, chromosomal, and collinearity analyses showed that these genes are evolutionarily conserved but structurally diverse. Promoter analysis detected abundant hormone and stress-related cis-acting elements, including 353 abscisic acid (ABA) responsive elements in 105 Qu-1-PsnNAC genes. Consistently, the enriched ACGT-core motif closely matched ABA-related bZIP motifs, including AREB3, ABF2, and ABI5, further linking Qu-1-PsnNAC promoters to ABA-responsive regulatory networks. Transcriptome analysis of Qu-1 suspension-cultured cells treated with 100 µM ABA at 0, 0.5, 4, 12, and 24 h revealed progressive transcriptional reprogramming over time. Thirty-two Qu-1-PsnNAC genes were significantly upregulated at one or more treatment time points. Integrated analysis of weighted gene coexpression network analysis (WGCNA) modules, Gene Ontology (GO) enrichment analysis, and promoter motif scanning suggested that a subset of Qu-1-PsnNACs and their predicted downstream genes may participate in ABA-responsive regulatory processes. Six key candidate genes were prioritized based on expression dynamics and network support. Quantitative real-time PCR (qRT-PCR) confirmed that these genes were inducible by 100 µM ABA at six time points (0, 0.5, 4, 12, 24, and 48 h) in buds, stems, leaves, and roots of wild-type (WT) P. simonii × P. nigra. Among them, Qu-1-PsnNAC41 and Qu-1-PsnNAC58 showed nuclear localization and transcriptional activation activity.

conclusionsThis study provides the first systematic characterization of the NAC gene family in the Qu-1 doubled haploid poplar system and establishes a framework for identifying ABA-associated NAC regulators and putative downstream targets. These findings expand our understanding of NAC-mediated ABA responses in poplar and provide valuable genetic resources for stress-tolerance improvement in tree breeding.