Cloning and Functional Analysis of the RiACO1 Gene in Raspberry

Red raspberry fruit is highly perishable, and raspberry plants are sensitive to drought and low-temperature stress because of their shallow root system, which limits production and postharvest utilization in cold regions. In this study, RiACO1 was cloned from red raspberry (‘Polka’) and analyzed by bioinformatics, subcellular localization, tissue-specific expression, heterologous overexpression in Arabidopsis thaliana, and transient overexpression in white-stage raspberry fruit. The full-length RiACO1 coding sequence was 963 bp and encoded a 320-amino-acid protein that localized to the cytoplasm and nucleus. RiACO1-overexpressing Arabidopsis lines showed higher survival rates under drought and low-temperature stress, accompanied by increased proline content, chlorophyll retention, and antioxidant enzyme activities, as well as reduced Malondialdehyde (MDA) and Reactive Oxygen Species (ROS) accumulation. In raspberry fruit, transient RiACO1 overexpression increased RiACO1 transcript levels, ACO activity, and ethylene production and was associated with accelerated softening, anthocyanin accumulation, and chlorophyll degradation. These results indicate that RiACO1 is involved in ethylene-associated fruit ripening and may contribute to abiotic-stress responses; however, its direct breeding value in raspberry requires further validation through stable raspberry transformation or targeted loss-of-function approaches.