Protein Phosphorylation Orchestrates Acclimations of Arabidopsis Plants to Environmental pH

Abstract

Environment pH (pHe) is a key parameter that dictates a surfeit of conditions critical to plant survival and fitness. To elucidate the mechanisms that recalibrate cytoplasmic and apoplastic pH homeostasis, we conducted a comprehensive proteomic/phosphoproteomic inventory of plants subjected to transient exposure to acidic or alkaline pH, an approach that covered the majority of protein-coding genes of the model plant Arabidopsis thaliana. Our survey revealed a large set so far undocumented pHe-dependent and potentially pH-specific phospho-sites, indicative of extensive post-translational regulation of proteins involved in the acclimation to pHe. Changes in pHe altered both electrogenic H+ pumping via P-type ATPases and H+/anion co-transport processes, leading to massively altered net trans-plasma membrane translocation of H+ ions. In pH 7.5 plants, transport (but not the assimilation) of nitrogen via NRT2-type nitrate and AMT1- type ammonium transporters was induced, conceivably to increase the cytosolic H+ concentration. Exposure to acidic pH resulted in a marked repression of primary root elongation. No such cessation was observed in nrt2.1 mutants, suggesting a role of NRT2.1 in negatively regulating root growth in response to acidic pH. Sequestration of iron into the vacuole via phosphorylation and abundance of the vacuolar iron transporter VTL5 was inversely regulated in response to high and low pHe, presumptively in anticipation of changes in iron availability associated with alterations of pHe in the soil. A pH-dependent phospho-switch was also observed for the ABC transporter PDR7, suggesting changes in activity and, possibly, substrate specificity. Unexpectedly, the effect of pHe was not restricted to roots and provoked pronounced changes in the leaf proteome. In both roots and shoots, the plant-specific TPLATE complex components AtEH1 and AtEH2 - essential for clathrin-mediated endocytosis - were differentially phosphorylated at multiple sites in response to pHe, indicating that the endocytic cargo protein trafficking is orchestrated by pHe.