Environmental temperature significantly affects plant growth and development, particularly flower development. In pepper (Capsicum annuum), the molecular mechanisms underlying temperature-mediated floral organ development remain unclear. Gibberellins (GAs) are key plant hormones regulating growth and development, including flower development, and the CaGA20ox gene family may play a crucial role in this process due to its involvement in GA biosynthesis. In this study, we comprehensively analyzed the CaGA20ox gene family across six pepper genomes (‘Zhangshugang’, ‘Zunla’, ‘Chiltepin’, ‘CM334’, ‘Ca59’, and ‘T2T’) to explore their roles in flower development and temperature stress response, identifying five to six genes per genome. These genes exhibited distinct expression patterns across different tissues and developmental stages, with some members showing higher expression in specific floral organs, particularly pistils. Our results revealed that temperature significantly impacts pepper flower development and GA content, with lower temperatures enhancing antioxidant capacity and increasing GA levels. Specifically, the expression levels of four CazGA20ox genes (CazGA20ox1, CazGA20ox2, CazGA20ox4, and CazGA20ox6) were significantly influenced by temperature changes. Our systematic analysis of the role of the CaGA20ox gene family in temperature-mediated pepper flower development provides a foundation for further studies on the molecular mechanisms as well as the development of improved pepper varieties.

AT-hook motif nuclear-localized (AHL) genes play critical roles in chromatin remodeling and gene transcription regulation, profoundly influencing plant growth, development, and stress responses. While AHL genes have been extensively characterized in multiple plant species, their biological functions in pepper (Capsicum annuum L.) remain largely uncharacterized. In this study, we identified 45 CaAHL genes in the pepper genome through bioinformatics approaches. Comprehensive analyses were conducted to examine their chromosomal distribution, phylogenetic relationships, and the structural and functional features of their encoded proteins. Phylogenetic clustering classified the CaAHL proteins into six distinct subgroups. Transcriptome profiling revealed widespread expression of CaAHL genes across diverse tissues—including roots, stems, leaves, flowers, seeds, pericarp, placenta, and fruits—at various developmental stages. Quantitative real-time PCR further demonstrated that CaAHL1, CaAHL33, and CaAHL23 exhibited consistently high expression throughout flower bud development, whereas CaAHL36 showed preferential upregulation at early bud development stages. Expression profiling under hormone treatments and abiotic stresses indicated that CaAHL36 and CaAHL23 are auxin-inducible but are repressed by ABA, cold, heat, salt, and drought stress. Subcellular localization assays in Nicotiana benthamiana leaf epidermal cells showed that both CaAHL36 and CaAHL23 were predominantly localized in the nucleus, with faint expression also detected in the cytoplasm. Collectively, this study provides foundational insights into the CaAHL gene family, laying the groundwork for future functional investigations of these genes in pepper.

The AT-hook motif nuclear-localized (AHL) genes play crucial roles in chromatin remodeling and regulation of gene transcription, significantly influencing biological processes such as plant growth, development, and stress responses. Although AHL genes have been extensively studied in various plants, their biological roles in pepper (Capsicum annuum L.) remain unknown. In this study, 45 AHL genes were identified in the pepper genome through bioinformatics approaches. These genes were analyzed for their chromosomal distribution, phylogenetic relationships, and the structural and functional characteristics of their encoded proteins. Phylogenetic analysis categorized the CaAHL proteins into four subgroups. Transcriptome data revealed that CaAHL genes are expressed across various tissues, including roots, stems, flowers, seeds, and fruits, at different growth stages. Subcellular localization experiments confirmed that CaAHL23 predominantly localizes to the cell nucleus, consistent with the localization of its homolog TEK in Arabidopsis thaliana. A yeast two-hybrid assay revealed that CaAHL23 binds to the promoter of CaCYP703A2, which is the homolog of the genic male sterility gene SlCYP703A2 in tomato. These findings indicate a potential link between CaAHL23 and genic male fertility in pepper. In summary, this study provides valuable insights that establish a foundation for future functional studies of CaAHL genes in pepper.