Silencing Sl-EBF1 and Sl-EBF2 expression causes constitutive ethylene response phenotype, accelerated plant senescence, and fruit ripening in tomato

The hormone ethylene regulates a wide range of plant developmental processes and EBF (EIN3-binding F-box) proteins were shown to negatively regulate the ethylene signalling pathway via mediating the degradation of EIN3/EIL proteins. The present study reports on the identification of two tomato F-box genes, Sl-EBF1 and Sl-EBF2 from the EBF subfamily. The two genes display contrasting expression patterns in reproductive and vegetative tissues and in response to ethylene and auxin treatment. Sl-EBF1 and Sl-EBF2 genes are actively regulated at crucial stages in the development of the reproductive organs. Their dynamic expression in flowers during bud-to-anthesis and anthesis-to-post-anthesis transitions, and at the onset of fruit ripening, suggests their role in situations where ethylene is required for stimulating flower opening and triggering fruit ripening. VIGS-mediated silencing of a single tomato EBF gene uncovered a compensation mechanism that tends to maintain a threshold level of Sl-EBF expression via enhancing the expression of the second Sl-EBF gene. In line with this compensation, tomato plants silenced for either of the Sl-EBF genes were indistinguishable from control plants, indicating functional redundancy among Sl-EBF genes. By contrast, co-silencing of both Sl-EBFs resulted in ethylene-associated phenotypes. While reports on EBF genes to date have focused on their role in modulating ethylene responses in Arabidopsis, the present study uncovered their role in regulating crucial stages of flower and fruit development in tomato. The data support the hypothesis that protein degradation via the ubiquitin/26S proteasome pathway is a control point of fruit ripening and open new leads for engineering fruit quality.