Abscisic acid (ABA) plays a vital role in coordinating physiological processes during fresh fruit ripening. ABA can bind to ABA receptors which interacts and inhibits their co-receptors type 2C phosphatases (PP2Cs). However, the dissected mechanism of PP2C during fruit ripening is unclear. In this study, we identify the role of SlPP2C3, a tomato type 2C phosphatase, as a negative regulator of ABA signaling and fruit ripening. SlPP2C3 selectively interacted with monomeric ABA receptors and SlSnRK2.8 kinase in both yeast and tobacco epidermal cells. Expressions of SlPP2C3 were observed in all tissues, and it negatively correlated with the fruit ripening which was induced by exogenous ABA. Tomato plants with suppressed SlPP2C3 expression exhibited enhanced sensitivity to ABA, while SlPP2C3 over-expressed plants were less sensitive to ABA. Meaningfully, lack of SlPP2C3 expression causes the acceleration of fruit ripening onset via the alternation of ABA signaling activity, and the fruit gloss is affected by the changes of outer epidermis structure. RNA-seq analysis found significant different expression of cuticle-related genes in pericarp between wild-type and SlPP2C3 suppressed lines. Taken together, our finding demonstrate that SlPP2C3 plays an important role in the regulation of fruit ripening and fruit appearance quality in tomato. 

RNA-seq

Total RNA was extracted from the skin of WT and SlPP2C3-RNAi4 fruits at the mature green stage. RNA degradation and contamination were monitored on 1% agarose gels. RNA integrity was assessed by an RNA 6000 Pico Assay Kit and a Bioanalyzer 2100 system (Agilent Technologies). The results were shown in Fig. S3. RNA (3 μg per sample) was used for mRNA purification and library construction with a Truseq RNA Sample Prep Kit (Illumina, CA, USA) following the manufacturer’s instructions. Samples were sequenced on an Illumina HiSeq 2000 platform (Illumina). Each sample yielded more than 6 Gb of data.

Raw data in fastq format were firstly processed using in-house perl scripts. Clean data was obtained by removing reads containing adapters, and poly-N and low-quality reads from raw data. Filtered RNA-seq clean reads were aligned to the reference tomato genome (release SL2.50) using TopHat v2.0.13 (Kim et al., 2013). HTSeq v0.5.3 (https://htseq.readthedocs.io/en/master/) was used to count the number of reads mapped to each gene, and reads per kilobase of transcript per million mapped reads (RPKM) values were calculated based on the length of the gene and the number of reads mapped. Differential expression analysis was performed using the DEGSeq R package (1.12.0) and Cufflinks (Trapnell et al., 2012). Corrected p-values of 0.001 and log2 (fold change) ³1 were set as the thresholds for significant differential expression.

Highlight: Tomato ABA co-receptor type 2C phosphatase (SlPP2C3), which acts as an ABA signalling negative regulator, affects tomato fruit ripening onset and fruit glossiness.