Aquaporins facilitate hydrogen peroxide entry into guard cells to mediate ABA- and pathogen-triggered stomatal closure.

Abstract

Stomatal movements are crucial for the control of plant water status and protection against pathogens. Assays on epidermal peels revealed that, similar to abscisic acid (ABA), pathogen-associated molecular pattern (PAMP) flg22 requires the <i>At</i>PIP2;1 aquaporin to induce stomatal closure. Flg22 also induced an increase in osmotic water permeability (<i>P</i>_f) of guard cell protoplasts through activation of <i>At</i>PIP2;1. The use of HyPer, a genetic probe for intracellular hydrogen peroxide (H₂O₂), revealed that both ABA and flg22 triggered an accumulation of H₂O₂ in wild-type but not <i>pip2;1</i> guard cells. Pretreatment of guard cells with flg22 or ABA facilitated the influx of exogenous H₂O₂ Brassinosteroid insensitive 1-associated receptor kinase 1 (BAK1) and open stomata 1 (OST1)/Snf1-related protein kinase 2.6 (SnRK2.6) were both necessary to flg22-induced <i>P</i>_f and both phosphorylated <i>At</i>PIP2;1 on Ser121 in vitro. Accumulation of H₂O₂ and stomatal closure as induced by flg22 was restored in <i>pip2;1</i> guard cells by a phosphomimetic form (Ser121Asp) but not by a phosphodeficient form (Ser121Ala) of <i>At</i>PIP2;1. We propose a mechanism whereby phosphorylation of <i>At</i>PIP2;1 Ser121 by BAK1 and/or OST1 is triggered in response to flg22 to activate its water and H₂O₂ transport activities. This work establishes a signaling role of plasma membrane aquaporins in guard cells and potentially in other cellular context involving H₂O₂ signaling.