Identification of a conserved set of cytokinin-responsive genes expressed in the fruits of Prunus persica

Ethylene plays an essential role in the ripening of peach, a climacteric stone fruit. However, several studies have shown that other phytohormones may alter fruit organoleptic properties, nutritional value, and overall quality. Although cytokinin levels are highest during fruit set and early stages of fruit development, exogenous cytokinin application on the late stages of peach fruit development (post-lignification stage) alters the peach fruit ripening indicators and delays ripening. To better understand the molecular mechanisms by which cytokinin alters fruit ripening and fruit quality traits, we identified a conserved set of 26 peach cytokinin-regulated orthologs of the Arabidopsis cytokinin-regulated genes. All but two of these genes show expression in ripe peach fruits. The peach cytokinin-regulated genes map throughout the Prunus persica genome, located on all eight chromosome-scale pseudomolecules. However, eight of these orthologs are located on pseudomolecule 1, while five orthologs are on pseudomolecule 2. Gene Ontology enrichment analyses revealed that many of these genes are associated with metabolic processes. The region upstream of the transcription start site contained Cytokinin Response Elements (CKREs) in 77% (20/26) of these putative cytokinin-regulated genes. The cytokinin responsiveness of eight of these genes (PpeBPH1, PpeCKX4, PpeCPK28, PpeCYP18-2, PpeHSP90-7, PpeNRP2, PpeNIP1-1, and PpePCRP1) was confirmed by RT-qPCR analyses of ripe peach fruits treated exogenously with trans-zeatin for one hour. Therefore, by using a comparative genomic analysis between peach and Arabidopsis, we have identified a conserved set of cytokinin-regulated genes expressed in peach fruits that respond to exogenous application of trans-zeatin and, based upon the conserved cis-regulatory regions, are most likely regulated by the cytokinin Type-B Response Regulators.