Understanding How 'honeycrisp' Apples Maintain Crispness by Elucidating Molecular Mechanisms Involved in Softening and Ethylene Production

The ‘Honeycrisp’ (HC) apple is new cultivar that has become very popular due to its flavor and unique texture characteristics. A distinct feature of the apple is that it maintains firmness over extended periods of time, including in air, and thus differs from most cultivars.  Our objective is to elucidate the biochemical changes and molecular mechanisms that are responsible for the slow softening of HC compared to a rapidly softening cultivar ‘McIntosh’ (MC). Fruits from both cultivars were picked during the normal harvest period and kept at 20 ^oC for two weeks. Internal ethylene concentrations (IECs) and flesh firmness of 5 fruit were measured every 3 d, and cortical tissue frozen in liquid nitrogen for further analysis.  From these samples, the tissue of three fruit for each sampling date/cultivar was used for RNA extraction. Primers were designed for various ACC synthases (ACSs), ACC oxidases (ACOs), polygalacturonase (PGs), ethylene receptor (ETRs and ERSs), xyloglucan endotransglucosylase (XTHs), expansions (EXPs), β-galactosidase (β-Gal), and arabinofuranosidase (AFase) genes, and used for quantitative real time RT-PCR (qPCR) analyses.  The results show that HC fruit kept at 20^oC had lower IECs, both at harvest and storage, than MC fruit.  Also, while MC fruit lost more than 33% of their initial firmness values, HC fruit remain almost as firm as at harvest. The qPCR results revealed that the expression of ACO1, ACO2, ACS1, ACS3, ERS1, ETR1, ETR12, ERF1, ERF2, and β-Gal genes by HC fruit were either higher than or equivalent in all dates to the expression level by MC fruits. It was evident that the expression of ACO and ACS genes in the earlier dates was much higher than MC fruits. However, the expression of AFase, PG, and EXP3 genes was significantly higher, at most dates, with MC fruit.