Mg-chelatase H subunit affects ABA signaling in stomatal guard cells, but is not an ABA receptor in Arabidopsis thaliana

Mg-chelatase H subunit (CHLH) is a multifunctional protein involved in chlorophyll synthesis, plastid-to-nucleus retrograde signaling, and ABA perception. However, whether CHLH acts as an actual ABA receptor remains controversial. Here we present evidence that CHLH affects ABA signaling in stomatal guard cells but is not itself an ABA receptor. We screened ethyl methanesulfonate-treated Arabidopsis thaliana plants with a focus on stomatal aperture-dependent water loss in detached leaves and isolated a rapid transpiration in detached leaves 1 (rtl1) mutant that we identified as a novel missense mutant of CHLH. The rtl1 and CHLH RNAi plants showed phenotypes in which stomatal movements were insensitive to ABA, while the rtl1 phenotype showed normal sensitivity to ABA with respect to seed germination and root growth. ABA-binding analyses using ³H-labeled ABA revealed that recombinant CHLH did not bind ABA, but recombinant pyrabactin resistance 1, a reliable ABA receptor used as a control, showed specific binding. Moreover, we found that the rtl1 mutant showed ABA-induced stomatal closure when a high concentration of extracellular Ca²⁺ was present and that a knockout mutant of Mg-chelatase I subunit (chli1) showed the same ABA-insensitive phenotype as rtl1. These results suggest that the Mg-chelatase complex as a whole affects the ABA-signaling pathway for stomatal movements.