The tomato zeaxanthin epoxidase-independent ABA biosynthetic pathway contributes to the synthesis of the root suberized exodermis

Recent studies have identified a zeaxanthin epoxidase-independent ABA biosynthetic pathway in Arabidopsis thaliana L., which operates through an apocarotenoid metabolic network conserved in the monocot rice but not yet explored in dicot crops. We investigated the presence of this alternative pathway in tomato ( Solanum lycopersicum L.), a carotenoid- and apocarotenoid-rich crop system. In exogenous application studies, we demonstrated that both β-apo-11-carotenal and its hydroxylated derivative, 3-OH-β-apo-11-carotenal, effectively inhibit seed germination and seedling establishment in wild-type tomato. However, the flacca mutant, which is impaired at the last step of ABA biosynthesis, did not show this response. Similarly, inhibition of hypocotyl and root growth by β-apo-11-carotenals was less pronounced in flacca than in WT, suggesting that these effects involve feeding ABA biosynthesis. Indeed, deuterated β-apo-11-carotenal and 3-OH-β-apo-11-carotenal were converted into deuterated ABA. Moreover, 6-hour treatment with 3-OH-β-apo-11-carotenal increased ABA levels in roots and activated ABA signaling. The transcriptional response, analyzed by RNA-seq, showed strong overlap with that of ABA treatment, although a subset of apocarotenoid-specific, ABA-independent-regulated genes was also identified. Notably, 3-OH-β-apo-11-carotenal treatment recapitulated ABA's induction of drought-response pathways, including comprehensive upregulation of the entire suberin biosynthetic machinery. This coordinated transcriptional activation enhanced suberin deposition in root exodermis—a critical adaptation for drought tolerance in tomato. Our findings demonstrate that 3-OH-β-apo-11-carotenal functions as a precursor in ABA biosynthesis, activates both ABA-dependent and independent signaling pathways, and promotes the formation of the root suberin barrier through systemic upregulation of its biosynthetic pathway. These results reveal the biological significance of the alternative pathway through suberin-mediated root protection.