ABA Biosynthesis- and Signaling-Related Gene Expression Differences between Sweet Cherry Fruits Suggest Attenuation of ABA Pathway in Bicolored Cultivars.

Fruit development involves exocarp color evolution. However, signals that control this process are still elusive. Differences between dark-red and bicolored sweet cherry cultivars rely on MYB factor gene mutations. Color evolution in bicolored fruits only occurs on the face receiving sunlight, suggesting the perception or response to color-inducing signals is affected. These color differences may be related to synthesis, perception or response to abscisic acid (ABA), a phytohormone responsible for non-climacteric fruit coloring. This work aimed to determine the involvement of ABA in the coloring process of color-contrasting varieties. Several phenolic accumulation patterns differed between bicolored 'Royal Rainier' and dark-red 'Lapins'. Transcript abundance of ABA biosynthetic genes (PavPSY, PavZEP and PavNCED1) decreased dramatically from the Pink to Red stage in 'Royal Rainier' but increased in 'Lapins', which correlated with a higher ABA content in this dark-red cultivar. Transcripts coding for ABA signaling (PavPP2Cs, PavSnRKs and PavMYB44.1) were almost undetectable at the Red stage in 'Royal Rainier'. Field trials revealed that 'Royal Rainier' color development was insensitive to exogenous ABA, whereas it increased in 'Lapins'. Furthermore, ABA treatment only increased transcript levels of signaling genes in 'Lapins'. Further studies may address if the ABA pathway is attenuated in bicolor cultivars.

Differential Phenolic Compounds and Hormone Accumulation Patterns between Early- and Mid-Maturing Sweet Cherry (Prunus avium L.) Cultivars during Fruit Development and Ripening.

Color acquisition is one of the most distinctive features of fruit development and ripening processes. The color red is closely related to the accumulation of polyphenolic compounds, mainly anthocyanins, during sweet cherry fruit maturity. In non-climacteric fruit species like sweet cherry, the maturity process is mainly controlled by the phytohormone abscisic acid (ABA), though other hormones may also play a role. However, the coordinated stage-specific production of polyphenolic compounds and their relation with hormone content variations have not been studied in depth in sweet cherry fruits. To further understand the accumulation dynamics of these compounds (hormones and metabolites) during fruit development, two sweet cherry cultivars ("Lapins" and "Glenred") with contrasting maturity timing phenotypes were analyzed using targeted metabolic analysis. The ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) approach revealed that phenolic acids, flavonols, and flavan-3-ols accumulated mainly until the straw-yellow stage in the early-maturing cultivar, while accumulation was mainly at the green stage in the mid-maturing cultivar, suggesting a cultivar-dependent stage-specific production of secondary metabolites. In the mid-maturing cultivar, anthocyanins were detected only from the red stage onward, whereas detection began at the pink stage in the early-maturing cultivar. ABA negatively correlated (p-value < 0.05) with the flavonols and flavan-3-ols in both cultivars. ABA and anthocyanin content increased at the same time in the early-season cultivar. In contrast, anthocyanins accumulated and the pink color initiation started several days after the ABA increase in the mid-maturing cultivar. Differential accumulation patterns of GA4, a ripening antagonizing hormone, between the cultivars could explain this difference. These results showed that both red-colored cultivars presented different accumulation dynamics of phenolic compounds and plant hormones during fruit development, suggesting underlying differences in the sweet cherry fruit color evolution.

RNAseq reveals different transcriptomic responses to GA3 in early and midseason varieties before ripening initiation in sweet cherry fruits.

Gibberellin (GA) negatively affects color evolution and other ripening-related processes in non-climacteric fruits. The bioactive GA, gibberellic acid (GA3), is commonly applied at the light green-to-straw yellow transition to increase firmness and delay ripening in sweet cherry (Prunus avium L.), though causing different effects depending on the variety. Recently, we reported that GA3 delayed the IAD parameter (a ripening index) in a mid-season variety, whereas GA3 did not delay IAD but reduced it at ripeness in an early-season variety. To further explore this contrasting behavior between varieties, we analyzed the transcriptomic responses to GA3 applied on two sweet cherry varieties with different maturity time phenotypes. At harvest, GA3 produced fruits with less color in both varieties. Similar to our previous report, GA3 delayed fruit color initiation and IAD only in the mid-season variety and reduced IAD at harvest only in the early-season variety. RNA-seq analysis of control- and GA3-treated fruits revealed that ripening-related categories were overrepresented in the early-season variety, including 'photosynthesis' and 'auxin response'. GA3 also changed the expression of carotenoid and abscisic acid (ABA) biosynthetic genes in this variety. In contrast, overrepresented categories in the mid-season variety were mainly related to metabolic processes. In this variety, some PP2Cs putative genes were positively regulated by GA3, which are negative regulators of ABA responses, and MYB44-like genes (putative repressors of PP2Cs expression) were downregulated. These results show that GA3 differentially modulates the transcriptome at the onset of ripening in a variety-dependent manner and suggest that GA3 impairs ripening through the modification of ripening associated gene expression only in the early-season variety; whereas in the mid-season variety, control of the ripening timing may occur through the PP2C gene expression regulation. This work contributes to the understanding of the role of GA in non-climacteric fruit ripening.

Gibberellic Acid Modifies the Transcript Abundance of ABA Pathway Orthologs and Modulates Sweet Cherry (Prunus avium) Fruit Ripening in Early- and Mid-Season Varieties.

Several phytohormones modulate ripening in non-climacteric fruits, which is triggered by abscisic acid (ABA). Gibberellins (GAs) are present during the onset of ripening in sweet cherry fruits, and exogenous gibberellic acid (GA3) application delays ripening, though this effect is variety-dependent. Although an ABA accumulation delay has been reported following GA3 treatment, the mechanism by which GA modulates this process has not been investigated at the molecular level in sweet cherry. Therefore, the aim of this work is to analyze the effect of GA3 on the fruit ripening process and the transcript levels of ABA pathway orthologs in two varieties having different maturity time phenotypes. The early-season variety had a rapid transition from yellow to pink fruit color, whereas pink color initiation took longer in the mid-season variety. GA3 increased the proportion of lighter colored fruits at ripeness in both varieties, but it produced a delay in IAD-a ripening index-only in the mid-season variety. This delay was accompanied by an increased transcript abundance of PavPP2Cs, which are putative negative regulators of the ABA pathway. On the other hand, the early-season variety had increased expression of PavCYP707A2-a putative ABA catabolic gene-, and reduced transcript levels of PavPP2Cs and SnRK2s after the GA3 treatment. Together these results show that GA modulates fruit ripening, exerting its action in part by interacting with the ABA pathway in sweet cherry.