Genome-Wide Identification, Characterization of the NIN-Like Protein Gene Family in Phoebe bournei and Its Heat-Responsive Expression Patterns

Abstract NIN-like proteins (NLPs) serve as transcription factors specific to plants that serve a central function in signaling and stress response in terms of nitrogen; however, their contribution to heat tolerance in woody plants remains poorly understood. Here, we identified 6 PbNLP genes in Phoebe bournei (Hemsl.) Yen C. Yang, an endangered tree species, and characterized their evolutionary relationships as well as gene and promoter elements, predicted protein structures and interactions, functional enrichment, tissue-expression profiles, and heat-responsive dynamic expression patterns with physiological indicators. The 6 genes were distributed among three of the four phylogenetic groups, and segmental duplication appeared to be the principal driver of family expansion. All PbNLP proteins possessed intact RWP-RK and PB1 domains, and their promoters were enriched in stress and hormone-responsive cis-elements. Secondary and tertiary structure predictions indicated a predominance of random coils, an absence of predicted beta-turns, and a conserved architecture comprising a compact core and flexible flanking regions. Protein interaction analysis predicted associations with several regulatory partners, particularly ROPGEF14, with PbNLP3 showing the highest network connectivity. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment showed the PbNLP family Complete stress responses via hormone‑signaling pathways. Transcript abundance was generally higher in leaves and bark than in xylem. Importantly, under heat stress, PbNLP genes showed expression divergence and most of them were positively correlated with one another, while differing substantially in their temporal expression trends, which were divided into two major response types. The PbNLP3 showed negative correlation with other genes and higest expression level. Analysis of heatmap revealed PbNLP5 exhibited the strongest correlations with photosynthetic physiological traits. Collectively, these findings identify PbNLP3 and PbNLP5 as candidate gene for further investigation of a potential crosstalk between nitrogen signaling and heat stress responses in P. bournei and provide potential targets for breeding heat-tolerant trees.