Genome-wide identification of RCC1 gene family in pepper (Capsicum annuum L.) and functional analysis of CaRCC1-16 under heat stress.

key messageCaRCC1-16 confers thermotolerance in pepper by interacting with stress-related proteins, enhancing photosynthetic efficiency, regulating stomatal activity, and modulating the expression of stress-responsive genes. Pepper (Capsicum annuum L.) is an economically important crop with high medicinal and biopesticidal properties that faces severe yield losses due to abiotic stresses. The regulator of chromosomal condensation 1 (RCC1) family protein is known to play important roles in plant growth, development, and stress responses. While several RCC1 members have been functionally characterized in Arabidopsis and other species, their role in pepper remains largely unexplored. In this study, we identified 28 CaRCC1 genes in pepper genome, unevenly distributed across 11 chromosomes. Phylogenetic characterization divided these genes into six groups, with members within each group sharing conserved gene structure and protein motif/domain structure. Promoter analysis revealed a high density of cis-acting elements associated with growth, hormone signaling, and stress responses. Furthermore, transcript profiling ABA and various abiotic stresses (heat, cold, drought, and salinity) revealed differential expression patterns, with CaRCC1-16 showing prominent transcription induction. Subcellular localization confirmed that CaRCC1-16 localized in the nucleus. Bimolecular fluorescence complementation analysis shows the interaction of CaRCC1-16 with ATpase, E2 18 and ETIF3 in the cytoplasm. Functional analyses of CaRCC1-16 demonstrate its role as a positive regulator of thermotolerance in pepper. Overexpression of CaRCC1-16 in pepper enhances heat tolerance by improving photosynthetic efficiency, regulating stomatal activity, reducing lipid peroxidation, and mitigating reactive oxygen species accumulation. In contrast, silencing of CaRCC1-16 results in increased heat sensitivity, overall, these findings elucidate the evolutionary history of the CaRCC1 family and, insights into the molecular mechanisms underlying pepper response to high temperature stress and, more significantly, define a previously unknown and important role for CaRCC1-16 in mediating the heat stress response through a novel protein interaction network. Contributing to our understanding of the biological functions of the RCC1 gene family in plant abiotic stress interaction, provides a foundation for crop improvement strategies.