Arabidopsis ALIX Regulates Stomatal Aperture and Turnover of Abscisic Acid Receptors

[EN] The plant endosomal trafficking pathway controls the abundance of membrane-associated soluble proteins, as shown for abscisic acid (ABA) receptors of the PYRABACTIN RESISTANCE1/PYR1-LIKE/REGULATORY COMPONENTS OF ABA RECEPTORS (PYR/PYL/RCAR) family. ABA receptor targeting for vacuolar degradation occurs through the late endosome route and depends on FYVE DOMAIN PROTEIN REQUIRED FOR ENDOSOMAL SORTING1 (FYVE1) and VACUOLAR PROTEIN SORTING23A (VPS23A), components of the ENDOSOMAL SORTING COMPLEX REQUIRED FOR TRANSPORT-I (ESCRT-I) complexes. FYVE1 and VPS23A interact with ALG-2 INTERACTING PROTEIN-X (ALIX), an ESCRT-III-associated protein, although the functional relevance of such interactions and their consequences in cargo sorting are unknown. In this study we show that Arabidopsis (Arabidopsis thaliana) ALIX directly binds to ABA receptors in late endosomes, promoting their degradation. Impaired ALIX function leads to altered endosomal localization and increased accumulation of ABA receptors. In line with this activity, partial loss-of-function alix-1 mutants display ABA hypersensitivity during growth and stomatal closure, unveiling a role for the ESCRT machinery in the control of water loss through stomata. ABA-hypersensitive responses are suppressed in alix-1 plants impaired in PYR/PYL/RCAR activity, in accordance with ALIX affecting ABA responses primarily by controlling ABA receptor stability. ALIX-1 mutant protein displays reduced interaction with VPS23A and ABA receptors, providing a molecular basis for ABA hypersensitivity in alix-1 mutants. Our findings unveil a negative feedback mechanism triggered by ABA that acts via ALIX to control the accumulation of specific PYR/PYL/RCAR receptors.This research was supported by the Ministerio de Economia y Competitividad (MINECO) and AEI/FEDER/EU (grants BIO2013-46539-R and BIO2016-80551-R to V.R., and BIO2017-82503-R to P.L.R.); by the Spanish Ministry of Education FPI "Severo Ochoa" fellowships (BES2015071820 to M.G.-L.); by the European Molecular Biology Organization (EMBO) Short-Term programs (ASTF 7678-2016); by the Agence Nationale de Recherche REGLISSE project (ANR-13-ADAP-0008 to L.C.); by an Agencia Espanola de Cooperacion Internacional (AECID) fellowship to D.A.E.-M.; by the Spanish Ministry of Education "Salvador de Madariaga" grant (PRX15/00493 to V.R.); and by Generalitat Valenciana "Programa VALi+d" (APOSTD/2017/039 to B.B.P.). We are thankful to Erika Isono for helpful discussion of the article. We thank Qi Xie for the anti-PYL4 antibody, and Oscar Lorenzo for cra1 seeds. We also thank the confocal microscopy, in vitro plant culture, and greenhouse facilities at the CNB-CSIC for technical support.Garcia, M.; Cuyas, L.; Abd El-Moneim, D.; Rodriguez, L.; Belda Palazón, B.; Sanchez, E.; Fernandez, Y.... (2019). Arabidopsis ALIX Regulates Stomatal Aperture and Turnover of Abscisic Acid Receptors. The Plant Cell. 31(10):2411-2429. https://doi.org/10.1105/tpc.19.00399S24112429311