The U-box E3 ubiquitin ligase (PUB) gene family plays an important role in regulating plant responses to abiotic stress. Poncirus trifoliata (trifoliate orange), a citrus rootstock with notable cold, drought, and salt tolerance, serves as an excellent model for studying stress-responsive genes. In this study, a total of 47 PUB genes (PtrPUBs) were identified in the trifoliate orange genome. Chromosomal distribution analysis indicated that PtrPUB genes were unevenly distributed across nine trifoliate orange chromosomes. Phylogenetic tree analysis indicated that 170 PUB proteins from trifoliate orange, Arabidopsis thaliana, and tomato were clustered into five subfamilies. Gene structure, conserved domain, and motif analyses revealed diverse exon–intron and motif organizations of PtrPUB genes, suggesting potential functional differentiation among PtrPUBs. Cis-acting analysis indicated that the promoters of PtrPUB genes harbor elements related to hormone signaling (ABA, MeJA), drought stress, and low-temperature responses. Transcriptomic data and qRT-PCR results suggested that PtrPUB genes are responsive to ABA and dehydration treatments. This study provides a foundation for understanding the functional roles of PUB genes in trifoliate orange and offers insights for improving stress tolerance in citrus breeding programs.

In this study, we identified 34 HD-ZIP genes at the genome-wide level in P. trifoliata (Poncirus trifoliata (L.) Raf.), and conducted comprehensive bioinformatic analyses. The expression profiles of PtrHB genes in response to six phytohormones (ABA, SA, JA, BR, ETH, GA) and two abiotic stresses (dehydration, cold) were further determined. Phylogenetic analysis assigned all PtrHB proteins to four distinct subfamilies (I–IV) with highly conserved domain compositions, in line with the established classification system in Arabidopsis thaliana. Segmental duplication was revealed as the primary driver of PtrHB family expansion, whereas three tandem duplication events were also detected. Promoter cis-element profiling revealed subfamily-specific distribution of hormone- and stress-responsive elements. Several candidate genes with prominent stimulus-specific responses were highlighted, including PtrHB32 (strongly induced by dehydration, cold and ABA), PtrHB15 (JA-responsive) and PtrHB16 (SA-responsive). Collectively, this study provides the first systematic genome-wide analysis of the HD-ZIP transcription factor family in P. trifoliata, and offers valuable insights into the evolutionary expansion and functional divergence of this family. Our results also lay a theoretical basis for functional validation of candidate stress-regulatory genes and for molecular breeding of stress-tolerant citrus rootstocks.