SlCNR regulates postharvest water loss and wax accumulation in tomato fruit and directly represses the transcription of very-long-chain (VLC) alkane biosynthesis-related genes SlCER1-2 and SlCER6

Cuticular waxes play a crucial role in the control of quality and storability of fruit and many transcription factors (TFs) contribute to fruit wax accumulation and affect the postharvest water loss. The tomato SBP-box protein Colorless Non-Ripening (SlCNR) is commonly investigated as a TF regulating fruit ripening, and its regulation role in mediating postharvest water loss remains to be unknown. Here, we found that fruit of SlCNR knockout (KO) mutants (KO-SlCNR#16 and KO-SlCNR#23) generated by CRISPR/Cas9 editing system exhibited decreased postharvest water loss, while fruit of the SlCNR-overexpressing (OE-SlCNR#21 and OE-SlCNR#24) displayed increased water loss compared to the wild type (WT). The overall wax accumulation increased by 17-30% in fruit of SlCNR KO mutants and decreased by 26-42% in OE-SlCNR#21 and OE-SlCNR#24, accompanied by the change of major components of waxes, such as n-alkanes, triterpenoids, and primary alcohol. RNA-seq analysis showed that the expression of numerous genes involved in wax biosynthesis were up-regulated in fruit of KO-SlCNR#23, including SlCER1-2, which encodes one part of the VLC alkane formation complex, and SlCER6, which encodes a beta-ketoacyl-coenzyme A synthase. Furthermore, we proved that SlCNR directly represses the transcription of wax biosynthetic genes SlCER1-2 and SlCER6 in vivo and in vitro, thereby negatively regulating wax accumulation. These findings expand our understanding in the genetic regulatory mechanisms governing ripe tomato fruit wax accumulation and present SlCNR as a novel target of wax accumulation and postharvest water loss.