Molecular physiology of ethylene during ripening of coffee fruits

Abstract

The quality of the coffee beans is directly associated with the ripening stages of the fruits at the time of harvest, which is often uneven due to the sequential flowering of the coffee tree, increasing the cost of production and generating a low-quality beverage. Some studies suggest that the coffee fruit is climacteric, indicating that ethylene plays an essential role in the ripening process. Growth regulators affect the speed of ripening of coffee fruits and improve harvest due to a higher synchronization of fruit ripening, which contributes to a beverage of higher quality. Tomato is a model species for the study of fleshy fruit development. Thus, Ethrel, Mathury, and 1-MCP were applied at different times to understand the physiological and genetic factors involved in regulating the ripening time of coffee fruits and the action of growth regulators in this process, and the molecular characterization of the ethylene biosynthesis (CaACO4) and signaling (CaERF1) genes using tomato (cv. Micro-Tom) as a heterologous species in natural variant introgressed lines, MT-rin and MT-nor. Coffee genomic regions upstream of each gene under study were fused with the coding region of GUS visible marker to characterize the promoter activity. GUS activity was evaluated in MT transgenic plants during the ripening of tomato fruits. The expression of genes that regulate ethylene biosynthesis and signaling, transcription factors that control ethylene production, genes induced by ethylene during fruit ripening, and the levels of ACC and activity of ACO were studied. The treatments with Ethrel, an ethylene inducer, showed better synchronization of fruit maturation, with approximately 70% of the fruits in the ripe stage (cherry) after harvest. Accordingly, such treatments with Ethrel showed increased expression of ethylene biosynthesis and signaling genes, CaACO4 and CaERF1, and ACC and ACO enzyme activity. Mathury, an ethylene inhibitor, caused an increase in the expression of genes from the APETALA2 (AP2) family and acted only on mature fruits, which decreased the amount of ACC and had a slight effect on the activity of the ACO enzyme. 1-MCP did not increase ACC levels and ACO activity, which suggested that there was no increase in endogenous ethylene levels. Thus, it is possible to affirm that 1-MCP can also be used as a growth regulator to delay the ripening of coffee fruits. Preliminary results with the transgenic lines MT-rin and MT-nor T0 plants overexpressing CaACO4 and CaERF1 genes showed that genes overexpressed, but the tomato fruits did not show a phenotype of ripening. The GUS test confirmed that CaERF1 has preferential activity in the fruit during ripening. The germination of MT-rin35S