Abscisic acid regulates Cl⁻ efflux via the ABI5-ZAT10-SLAH3 module in chloride-stressed Malus hupehensis

Abstract

The overload of Cl⁻ typically causes cell damage and death in plants, especially in Cl⁻-sensitive crops. Abscisic acid (ABA) is a stress-induced phytohormone that can alleviate chloride stress by reducing Cl⁻ accumulation; however, the mechanism is not clear. Here, we found that the application of ABA elevated Cl⁻ efflux from roots and reduced membrane damage and cell death in chloride-stressed Malus hupehensis. MhSLAH3, a homolog of the slow anion channel from M. hupehensis, encoded a channel controlling Cl⁻ efflux and was induced by both chloride and ABA. MhSLAH3 overexpression accelerated Cl⁻ efflux, which enhanced the tolerance of M. hupehensis to chloride stress, and retarded chloride-induced cell death. However, the suppression of MhSLAH3 partially offset the acceleration effect of ABA on Cl⁻ efflux. MhZAT10L was then identified as a C2H2-type transcription factor upstream of MhSLAH3, repressing MhSLAH3 transcription under chloride stress. The suppression of MhZAT10L accelerated Cl⁻ efflux by releasing suppressed MhSLAH3, but MhZAT10L overexpression counteracted the effects of ABA on Cl⁻ efflux. MhABI5 promoted Cl⁻ efflux mediated by MhSLAH3 due to induction by ABA and transcriptional repression of MhZAT10L, but this function of MhABI5 was reversed by MhZAT10L overexpression. The suppression of MhABI5 diminished the positive effects of ABA on Cl⁻ efflux and retarding cell death. Thus, ABA repressed MhZAT10L transcription by activating MhABI5, further releasing MhSLAH3 to accelerate Cl⁻ efflux. These findings provide a new evidence of ABA regulation of Cl⁻ efflux.

Acknowledgements

This work acknowledges the financial support from the National Natural Science Foundation of China (32172517), the National Key Research and Development Program of China (2019YFD1000103), and the Natural Science Foundation of Shandong Province (ZR2023MC003). We also thank the linguistic assistance provided by LetPub (www.letpub.com) in preparing this manuscript.

Author contributions

J.S.: Conceptualization, Investigation, Data curation, Formal analysis, Methodology, Visualization, Writing-original draft. J.Y.: Investigation, Methodology, Data curation. B.S.: Investigation. B.C.: Investigation. X.Z.: Investigation. H.W.: Investigation. Z.B.: Investigation, Resources. F.M.: Investigation, Resources. W.Z.: Conceptualization, Funding acquisition, Writing-review & editing, Resources. H.Y.: Project administration, Funding acquisition, Writing-review & editing, Resources.

Data availability

All data are available in the main text or the supplementary information.

Conflict of interest statement

The authors declare no conflicts of interest.

Supplementary data

Supplementary data is available at Horticulture Research online.

References

[1] Raven JA. Chloride: essential micronutrient and multifunctional beneficial ion. J Exp Bot. 2017; 68:359-67

[2] Broyer TC, Carlton A, Johnson C. et al. Chlorine-a micronutrient element for higher plants. Plant Physiol. 1954; 29:526-32

[3] Geilfus CM. Chloride in soil: from nutrient to soil pollutant. Environ Exp Bot. 2019; 157:299-309

[4] Sanders D. The salinity challenge. New Phytol. 2020; 225:1047-8

[5] Wani SH, Kumar V, Khare T. et al. Engineering salinity tolerance in plants: progress and prospects. Planta. 2020; 251:1-29

[6] Xu G, Magen H, Tarchitzky J. et al. Advances in chloride nutrition of plants. Adv Agron. 1999; 68:97-150

[7] Geilfus CM. Chloride: from nutrient to toxicant. Plant Cell Physiol. 2018; 59:877-86

[8] Isayenkov S. Physiological and molecular aspects of salt stress in plants. Cytol Genet. 2012; 46:302-18

[9] Zhang X, Franzisky BL, Eigner L. et al. Antagonism of chloride and nitrate inhibits nitrate reductase activity in chloride-stressed maize. Plant Growth Regul. 2021; 93:279-89

[10] Santos C, Malta MR, Gonçalves MGM. et al. Chloride applied via fertilizer affects plant nutrition and coffee quality. Plants(Bsael). 2023; 12:885

[11] Bañuls J, Primo-millo E. Effects of chloride and sodium on gas exchange parameters and water relations of citrus plants. Physiol Plant. 1992; 86:115-23

[12] Zhang W, Yang H, Su Q. et al. Response of nitric oxide and active oxygen of Malus hupehensis roots to sodium and chloride ion stress. Acta Hortic Sin. 2013; 40:2365

[13] Zhang W, Yang H, You S. et al. MhNCED3, a gene encoding 9-cis-epoxycarotenoid dioxygenase in Malus hupehensis Rehd., enhances plant tolerance to Cl^- stress by reducing Cl^- accumulation. Plant Physiol Biochem. 2015; 89:85-91

[14] Isayenkov SV, Maathuis FJM. Plant salinity stress: many unanswered questions remain. Front Plant Sci. 2019; 10:80

[15] Wu H, Li Z. The importance of Cl^- exclusion and vacuolar Cl^- sequestration: revisiting the role of Cl^- transport in plant salt tolerance. Front Plant Sci. 2019; 10:1418

[16] Kurusu T, Saito K, Horikoshi S. et al. An S-type anion channel SLAC1 is involved in cryptogein-induced ion fluxes and modulates hypersensitive responses in tobacco BY-2 cells. PLoS One. 2013; 8:e70623

[17] Song J, Han M, Zhu X. et al. MhCLC-c1, a Cl channel c homolog from Malus hupehensis, alleviates NaCl-induced cell death by inhibiting intracellular Cl^- accumulation. BMC Plant Biol. 2023; 23:306

[18] Hedrich R, Geiger D. Biology of SLAC1-type anion channels from nutrient uptake to stomatal closure. New Phytol. 2017; 216:46-61

[19] Cui Y, Li X, Liu R. et al. SLAH1 is involved in the long-distance transport of Cl^- from roots into shoots in the Cl^--tolerant xerophyte Pugionium cornutum under salt stress. Plant Soil. 2022; 479:631-48

[20] Qiu J, Henderson SW, Tester M. et al. SLAH1, a homologue of the slow type anion channel SLAC1, modulates shoot Cl^- accumulation and salt tolerance in Arabidopsis thaliana. J Exp Bot. 2016; 67:4495-505

[21] Cubero-Font P, Maierhofer T, Jaslan J. et al. Silent S-type Anion Channel subunit SLAH1 gates SLAH3 open for chloride root-to-shoot translocation. Curr Biol. 2016; 26:2213-20

[22] Jaborsky M, Maierhofer T, Olbrich A. et al. SLAH3-type anion channel expressed in poplar secretory epithelia operates in calcium kinase CPK-autonomous manner. New Phytol. 2016; 210:922-33

[23] Henderson SW, Baumann U, Blackmore DH. et al. Shoot chloride exclusion and salt tolerance in grapevine is associated with differential ion transporter expression in roots. BMC Plant Biol. 2014; 14:1-18

[24] Reed NR, Bonner BA. The effect of abscisic acid on the uptake of potassium and chloride into Avena coleoptile sections. Planta. 1974; 116:173-85

[25] Wang N, Wang X, Zhang H. et al. Early ABA-stimulated maintenance of Cl^- homeostasis by mepiquat chloride priming confers salt tolerance in cotton seeds. Crop J. 2021; 9:387-99

[26] Collin A, Daszkowska-Golec A, Szarejko I. Updates on the role of ABSCISIC ACID INSENSITIVE 5 (ABI5) and ABSCISIC ACID-RESPONSIVE ELEMENT BINDING FACTORS (ABFs) in ABA signaling in different developmental stages in plants. Cells. 2021; 10:1996

[27] Nakashima K, Fujita Y, Kanamori N. et al. Three Arabidopsis SnRK2 protein kinases, SRK2D/SnRK2.2, SRK2E/SnRK2.6/OST1 and SRK2I/SnRK2.3, involved in ABA signaling are essential for the control of seed development and dormancy. Plant Cell Physiol. 2009; 50:1345-63

[28] Liu W, Chen G, He M. et al. ABI5 promotes heat stress-induced chlorophyll degradation by modulating the stability of MYB44 in cucumber. Hortic Res. 2023; 10:uhad089

[29] Fan W, Xu JM, Wu P. et al. Alleviation by abscisic acid of Al toxicity in rice bean is not associated with citrate efflux but depends on ABI5-mediated signal transduction pathways. J Integr Plant Biol. 2019; 61:140-54

[30] Lei GJ, Zhu XF, Wang ZW. et al. Abscisic acid alleviates iron deficiency by promoting root iron reutilization and transport from root to shoot in Arabidopsis. Plant Cell Environ. 2014; 37:852-63

[31] Liu Y, Khan AR, Gan Y. C2H2 zinc finger proteins response to abiotic stress in plants. Int J Mol Sci. 2022; 23:2730

[32] He F, Li HG, Wang JJ. et al. PeSTZ1, a C2H2-type zinc finger transcription factor from Populus euphratica, enhances freezing tolerance through modulation of ROS scavenging by directly regulating PeAPX2. Plant Biotechnol J. 2019; 17:2169-83

[33] Li P, Wang M, Zhou Y. et al. Freezing transcriptome analysis showed that GhZAT10 regulates freezing tolerance through a partially CBF-dependent pathway in upland cotton (Gossypium hirsutum L.). Environ Exp Bot. 2023; 208:105263

[34] Li XL, Meng D, Li MJ. et al. Transcription factors MhDREB2A/MhZAT10 play a role in drought and cold stress response crosstalk in apple. Plant Physiol. 2023; 192:2203-20

[35] Yu L, Guo Z, Liu W. et al. Interaction between MdZAT10a-like and MdbHLH100 negatively regulates salt tolerance in apple (Malus domestica Borkh.). Environ Exp Bot. 2022; 200:104938

[36] Yang K, An JP, Li CY. et al. The apple C2H2-type zinc finger transcription factor MdZAT10 positively regulates JA-induced leaf senescence by interacting with MdBT2. Hortic Res. 2021; 8:159

[37] Yang K, Li CY, An JP. et al. The C2H2-type zinc finger transcription factor MdZAT10 negatively regulates drought tolerance in apple. Plant Physiol Biochem. 2021; 167:390-9

[38] Chen Y, Sun J, Wei Y. et al. PpZAT10 negatively regulates peach cold resistance predominantly mediated by inhibiting VIN activity. Postharvest Biol Technol. 2022; 190:111952

[39] Song J, Yan J, Sun B. et al. MhZAT10L positively modulates NaCl-induced cell death by triggering MhVPEγ transcription in Malus hupehensis. Environ Exp Bot. 2024; 223:105783

[40] Kumar N, Haldarb S, Saikia R. Root exudation as a strategy for plants to deal with salt stress: An updated review. Environ Exp Bot. 2023; 216:105518

[41] Bhatla SC, Lal MA. Concepts of Plant Water Relations. In: Bhala SC, Lal MA (eds.), Plant Physiology, Development and Metabolism. Singapore: Springer Nature Singapore, 2023, 3-23

[42] Scharwies JD, Dinneny JR. Water transport, perception, and response in plants. J Plant Res. 2019; 132:311-24

[43] Chen G, Wang L, Chen Q. et al. PbrSLAH 3 is a nitrate-selective anion channel which is modulated by calcium-dependent protein kinase 32 in pear. BMC Plant Biol. 2019; 19:190

[44] Lee SC, Lan W, Buchanan BB. et al. A protein kinase-phosphatase pair interacts with an ion channel to regulate ABA signaling in plant guard cells. Proc Natl Acad Sci USA. 2009; 106:21419-24

[45] Tavakkoli E, Fatehi F, Coventry S. et al. Additive effects of Na⁺ and Cl^- ions on barley growth under salinity stress. J Exp Bot. 2011; 62:2189-203

[46] Edgar RC. MUSCLE: a multiple sequence alignment method with reduced time and space complexity. Bioinformatics. 2004; 5:113

[47] Chen XS, Li TT, Zhou SL. et al. Transient expression of exogenous protein in tobacco leaves. Bio-protocol. 2018;e1010127

[48] Raffaele S, Bayer E, Lafarge D. et al. Remorin, a Solanaceae protein resident in membrane rafts and Plasmodesmata, ImpairsPotato virus XMovement. Plant Cell. 2009; 21:1541-55

[49] Zhang W, Wang Z, Song J. et al. Cd²⁺ uptake inhibited by MhNCED3 from Malus hupehensis alleviates Cd-induced cell death. Environ Exp Bot. 2019; 166:103802

[50] Song J, Yang F, Xun M. et al. Genome-wide identification and characterization of vacuolar processing enzyme gene family and diverse expression under stress in apple (Malus × domestic). Front Plant Sci. 2020; 11:626

[51] Chow CN, Lee TY, Hung YC. et al. PlantPAN3.0: a new and updated resource for reconstructing transcriptional regulatory networks from ChIP-seq experiments in plants. Nucleic Acids Res. 2019; 47:D1155-63