Unveiling the CpHsp and CpHsf Gene families in the zucchini genome: Evolution, structure, and stress-responsive expression

Abstract Proper protein folding and stability are vital for plant development and stress tolerance. Heat shock proteins (Hsps), regulated by heat shock transcription factors (Hsfs), play a central role in abiotic stress responses. Zucchini ( Cucurbita pepo ), a key member of the Cucurbitaceae family, is economically important and widely cultivated worldwide. In Turkey, Seyden F1 and Cordelia F1 are the predominant cultivars used for domestic consumption and export. In this study, a total of 464 C. pepo Hsp genes distributed across six subfamilies and 77 CpHsf genes were identified and analyzed using bioinformatics tools, representing the first comprehensive genome-wide identification and characterization of Hsp and Hsf gene families in C. pepo across two cultivars. Gene structure, conserved motifs, and phylogenetic relationships indicated strong evolutionary conservation, largely shaped by segmental duplications. Comparative genomic analyses revealed that soybean is the closest species based on shared orthologs. miRNA analyses highlighted miR395 and miR167 as key post-transcriptional regulators of CpHsp and CpHsf genes. Transcriptomic analyses revealed distinct tissue-specific and temporal expression patterns under normal and stress conditions. qRT-PCR validation in Seyden F1 and Cordelia F1 cultivars under heat, drought, and combined stress treatments identified key stress-responsive genes, CpHsp70-20 and CpHsf-68 , which were consistently upregulated across all stress conditions. Interestingly, CpHsp60-34 showed significant downregulation under heat stress in the Cordelia cultivar, highlighting genotype-specific variations between cultivars. The key marker genes and in silico interaction networks identified herein provide a robust resource for functional characterization and molecular breeding to improve stress resilience in zucchini.