Ethylene signaling is decisive for many plant developmental processes. Among these, control of senescence, abscission and fruit ripening are of fundamental relevance for global agriculture. Consequently, detailed knowledge of the signaling network along with the molecular processes of signal perception and transfer are expected to have high impact on future food production and agriculture. Recent advances in ethylene research have demonstrated that signaling of the plant hormone critically depends on the interaction of the ethylene receptor family with the NRAMP-like membrane protein ETHYLENE INSENSITIVE 2 (EIN2) at the ER membrane, phosphorylation-dependent proteolytic processing of ER-localized EIN2 and subsequent translocation of the cleaved EIN2 C-terminal polypeptide (EIN2-CEND) to the nucleus. EIN2 nuclear transport, but also interaction with the receptors sensing the ethylene signal, both, depend on a nuclear localization signal (NLS) located at the EIN2 C-terminus. Loss of the tight interaction between receptors and EIN2 affects ethylene signaling and impairs plant ethylene responses. Synthetic peptides derived from the NLS sequence interfere with the EIN2–receptor interaction and have utility in controlling plant ethylene responses such as ripening. Here, we report that a synthetic peptide (NOP-1) corresponding to the NLS motif of Arabidopsis EIN2 (aa 1262–1269) efficiently binds to tomato ethylene receptors LeETR4 and NR and delays ripening in the post-harvest phase when applied to the surface of sampled green fruits pre-harvest. In particular, degradation of chlorophylls was delayed by several days, as monitored by optical sensors and confirmed by analytical methods. Similarly, accumulation of β-carotene and lycopene in the fruit pulp after NOP-1 application was delayed, without having impact on the total pigment concentration in the completely ripe fruits. Likewise, the peptide had no negative effects on fruit quality. Our molecular and phenotypic studies reveal that peptide biologicals could contribute to the development of a novel family of ripening inhibitors and innovative ripening control in climacteric fruit.

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BACKGROUND Processes extending the shelf life of climacteric fruit play an important role in terms of a sustainable global food supply. In a previous study, a synthetic octapeptide (NOP‐1) was shown to inhibit the interaction between ethylene receptor (ETR) and ethylene insensitive‐2 (EIN2), and in consequence delay tomato ripening. We investigated for the first time the effect of NOP‐1 on inhibiting the ripening of apples (‘Golden Delicious’) during postharvest. RESULTS Using purified recombinant proteins from a bacterial expression system, we demonstrate here that EIN2 also interacts tightly (Kd = 136 ± 29 nmol L−1) with the corresponding apple ETR MdETR1. In line with previous binding studies on tomato ETRs, the ripening‐delaying peptide NOP‐1 clearly binds to the purified apple ETR. An NOP‐1 solution (1000 µmol L−1) was applied with a brush or microdispenser and compared with apples treated with 1‐methylcyclopropene (1‐MCP) (SmartFresh™, Agrofresh) applied as gaseous treatment or untreated control fruits. NOP‐1 inhibited colour development and chlorophyll degradation during shelf life. These effects were more pronounced with the brush application (surface film) than with microdroplets application (mimicking a sprayable formulation). NOP‐1 did not alter ethylene release or respiration rate, whereas 1‐MCP expectedly strongly suppressed both. There were no differences in quality parameters evaluated. CONCLUSION Our study shows that NOP‐1 binds to MdETR1 which results in delaying of ethylene‐dependent ripening developments of skin colour and chlorophyll. Besides application methods, possible reasons for the weak effect of NOP‐1 in comparison with previous tomato experiments could be different receptor affinity and penetration differences. © 2019 Society of Chemical Industry

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