Understanding the effect of calcium in kiwifruit ripening and establishment of early and late response mechanisms through a cross-omics approach

Calcium (Ca2+) is a secondary messenger that plays a pivotal role in kiwifruit ripening; however, the underlying mechanisms are still unclear. Herein, we characterize the physiological and molecular responses of kiwifruit to calcium nutrition and how these changes could influence fruit ripening. Our study addresses the response of kiwifruit to calcium in both the early (8 h after calcium application) and late (during ripening at room temperature following 3 months of cold storage) stages. Exogenously supplied calcium (2% CaCl2) by dipping induced an early endogenous calcium accumulation and stimulated intracellular Ca2+ signals from the inner to outer pericarp and in the vascular tissue. Calcium treatment delays kiwifruit ripening, as evidenced by the reduction of ethylene production and softening that is accompanied by altered levels of genes and proteins involved in ethylene signaling and cell wall structure. An immunomicroscopy approach at the early stage based on epitope distribution using cell wall antibodies such as LM19, JIM13 and LM30 showed that de-esterified homogalacturonans and arabinogalactan proteins were depressed in calcium-exposed fruit. The levels of primary metabolites were decreased, while several secondary metabolites, including epicatechin, catechin and procyanidin B2 were altered by calcium. Treatment with calcium has a profound long-term impact on kiwifruit’s transcriptome and proteome contributing to ripening changes. Particularly, the study uncovered an extensive transcriptomic regulation of ripening signaling, notably through MAPKs and hormone, by calcium. A wide diversity of transcription factors (TFs), particularly the AP2/ERF-ERF group, displayed distinct expression patterns at the early and late stages, providing potential targets for deciphering the initial TFs and late responses that are triggered by calcium. A calcium network of the genes-proteins-metabolites and the connected TFs was also created. Interlinked gene expression and protein accumulation analysis unveil that calcium elicits cysteine modifications, membrane/transporter activity, ascorbate homeostasis and ubiquitination signaling. From the obtained transcriptomic and proteomic data, we constructed a calcium-affected proteogenomic framework, highlighting a possible role for alternative splicing in ripening initiation. These findings provide new knowledge of the physiological processes, biochemical mechanisms, and networks regulated by calcium during kiwifruit ripening.