Pan-genome identification of the HSF gene family in pear and expression profiling of PbHSFs during postharvest cold storage

Heat shock transcription factors (HSFs) are key regulators of plant growth, development, and responses to environmental stresses. Although HSFs have been extensively studied in model plants and several fruit crops, their evolutionary patterns and stress-responsive functions in pear ( Pyrus spp.) particularly in the context of postharvest cold stress. ‘Huangguan’ pear is an economically important cultivar in China but highly susceptible to chilling injury during cold storage. The availability of multiple pear genome assemblies provides an opportunity for comprehensive pan-genome analysis of the pear HSF family and identification of cold-responsive PbHSF genes. In this study, we identified 527 HSF genes across 13 pear genomes and classified them into 21 core and 5 non-core orthogroups (OGs), revealing a highly conserved and core-dominated HSF family structure. Core OGs accounted for most HSF members in all genomes, indicating strong evolutionary conservation. Comparative duplication analysis showed that whole-genome duplication (WGD) – derived genes were significantly enriched in the HSF family, suggesting that WGD-derived gene retention has substantially contributed to HSF family expansion and diversification. Despite extensive expansion, representative HSF gene pairs from both core and non-core OGs exhibited strong purifying selection, suggesting these HSF members were retained during evolution. Transcriptome analysis during cold storage of ‘Huangguan’ pear revealed that several HSF genes belonging to core OGs responded strongly to low-temperature treatment. Among them, PbHSFC1 was identified as a candidate cold-responsive gene, showing pronounced induction under 0 °C storage and an expression pattern similar to that of the cold-signaling factor DREB1B . Promoter analysis further revealed that PbHSFC1 contains abundant ABA- and dehydration-responsive cis -elements, suggesting that its transcription may be regulated by ABA- and dehydration-related signaling pathways. This study provides a comprehensive pan-genome analysis of the pear HSF gene family and reveals the significant contribution of WGD-derived gene retention to HSF family expansion. Expression profiling under cold stress further suggested that PbHSFC1 may contribute to such responses. In particular, PbHSFC1 exhibited an expression pattern similar to DREB1B during cold storage. These findings provide a valuable foundation for future studies on cold tolerance in pears.