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Summary: Fruit firmness is an important trait in sweet cherry breeding because it directly positively influences fruit transportability, storage and shelf life. However, the underlying genes responsible and the molecular mechanisms that control fruit firmness remain unknown. In this study, we identified a candidate gene, PavSCPL, encoding a serine carboxypeptidase‐like protein with natural allelic variation, that controls fruit firmness in sweet cherry using map‐based cloning and functionally characterized PavSCPL during sweet cherry fruit softening. Genetic analysis revealed that fruit firmness in the 'Rainier' × 'Summit' F1 population was controlled by a single dominant gene. Bulked segregant analysis combined with fine mapping narrowed the candidate gene to a 473‐kb region (7418778–7 891 914 bp) on chromosome 6 which included 72 genes. The candidate gene PavSCPL, and a null allele harbouring a 5244‐bp insertion in the second exon that completely inactivated PavSCPL expression and resulted in the extra‐hard‐flesh phenotype, were identified by RNA‐sequencing analysis and gene cloning. Quantitative RT‐PCR analysis revealed that the PavSCPL expression level was increased with fruit softening. Virus‐induced gene silencing of PavSCPL enhanced fruit firmness and suppressed the activities of certain pectin‐degrading enzymes in the fruit. In addition, we developed functional molecular markers for PavSCPL and the Pavscpl5.2‐k allele that co‐segregated with the fruit firmness trait. Overall, this research identified a crucial functional gene for fruit firmness. The results provide insights into the genetic control and molecular mechanism of the fruit firmness trait and present useful molecular markers for molecular‐assisted breeding for fruit firmness in sweet cherry. [ABSTRACT FROM AUTHOR]

For sweet cherry, fruit size is one of the main targets in breeding programs owing to the high market value of larger fruits. KLUH/CYP78A5 is an important regulator of seed/fruit size in several plant species, but its molecular mechanism is largely unknown. In this study, we characterized the function of PavKLUH in the regulation of sweet cherry fruit size. The ectopic overexpression of PavKLUH in Arabidopsis increased the size of its siliques and seeds, whereas virus‐induced gene silencing of PavKLUH in sweet cherry significantly decreased fruit size by restricting mesocarp cell expansion. We screened out an AP2/ERF transcription factor containing a B3‐like domain, designated as PavRAV2, which was able to physically interact with PavKLUH promoter in a yeast one‐hybrid (Y1H) system. In Y1H assays, electrophoretic mobility shift assays, and dual‐luciferase reporter analyses, PavRAV2 directly bound to the promoter of PavKLUH in vitro and in vivo, and suppressed PavKLUH expression. Silencing of PavRAV2 resulted in enlarged fruit as a result of enhanced mesocarp cell expansion. Together, our results provide new insights into signaling pathways related to fruit size, and outline a possible mechanism for how the RAV transcription factor directly regulates CYP78A family members to influence fruit size and development. [ABSTRACT FROM AUTHOR]

The rapid softening of sweet cherry fruits during ripening results in the deterioration of fruit quality. However, few genes related to sweet cherry fruit ripening and softening have been identified, and the molecular regulatory mechanisms underlying this process are poorly understood. Here, we identified and functionally characterized PavNAC56, a NAC transcription factor that positively regulates sweet cherry fruit ripening and softening. Gene expression analyses showed that PavNAC56 was specifically and abundantly expressed in the fruit, and its transcript levels increased in response to abscisic acid (ABA). A subcellular localization analysis revealed that PavNAC56 is a nucleus‐localized protein. Virus‐induced gene silencing of PavNAC56 inhibited fruit ripening, enhanced fruit firmness, decreased the contents of ABA, anthocyanins, and soluble solids, and down‐regulated several fruit ripening‐related genes. Yeast one‐hybrid and dual‐luciferase assays showed that PavNAC56 directly binds to the promoters of several genes related to cell wall metabolism (PavPG2, PavEXPA4, PavPL18, and PavCEL8) and activates their expression. Overall, our findings show that PavNAC56 plays an indispensable role in controlling the ripening and softening of sweet cherry fruit and provides new insights into the regulatory mechanisms by which NAC transcription factors affect nonclimacteric fruit ripening and softening. [ABSTRACT FROM AUTHOR]