The stress hormone abscisic acid (ABA) induces expression of defence genes in many organs, modulates ion homeostasis and metabolism in guard cells, and inhibits germination and seedling growth. Concerning the latter effect, several mutants of Arabidopsis thaliana with improved capability for H⁺ efflux (wat1-1D, overexpression of AKT1 and ost2-1D) are less sensitive to inhibition by ABA than the wild type. This suggested that ABA could inhibit H⁺ efflux (H⁺-ATPase) and induce cytosolic acidification as a mechanism of growth inhibition. Measurements to test this hypothesis could not be done in germinating seeds and we used roots as the most convenient system. ABA inhibited the root plasma-membrane H⁺-ATPase measured in vitro (ATP hydrolysis by isolated vesicles) and in vivo (H⁺ efflux from seedling roots). This inhibition involved the core ABA signalling elements: PYR/PYL/RCAR ABA receptors, ABA-inhibited protein phosphatases (HAB1), and ABA-activated protein kinases (SnRK2.2 and SnRK2.3). Electrophysiological measurements in root epidermal cells indicated that ABA, acting through the PYR/PYL/RCAR receptors, induced membrane hyperpolarization (due to K⁺ efflux through the GORK channel) and cytosolic acidification. This acidification was not observed in the wat1-1D mutant. The mechanism of inhibition of the H⁺-ATPase by ABA and its effects on cytosolic pH and membrane potential in roots were different from those in guard cells. ABA did not affect the in vivo phosphorylation level of the known activating site (penultimate threonine) of H⁺-ATPase in roots, and SnRK2.2 phosphorylated in vitro the C-terminal regulatory domain of H⁺-ATPase while the guard-cell kinase SnRK2.6/OST1 did not.

Summary: Intracellular pH must be kept close to neutrality to be compatible with cellular functions, but the mechanisms of pH homeostasis and the responses to intracellular acidification are mostly unknown. In the plantArabidopsis thaliana, we found that intracellular acid stress generated by weak organic acids at normal external pH induces expression of several chaperone genes, includingROF2, which encodes a peptidyl‐prolylcis‐transisomerase of the FK506‐binding protein class. Loss of function ofROF2, and especially double mutation ofROF2and the closely related geneROF1, results in acid sensitivity. Over‐expression ofROF2confers tolerance to intracellular acidification by increasing proton extrusion from cells. The activation of the plasma membrane proton pump (H⁺‐ATPase) is indirect: over‐expression ofROF2activates K⁺uptake, causing depolarization of the plasma membrane, which activates the electrogenic H⁺pump. The depolarization ofROF2over‐expressing plants explains their tolerance to toxic cations such as lithium, norspermidine and hygromycin B, whose uptake is driven by the membrane potential. AsROF2induction and intracellular acidification are common consequences of many stresses, this mechanism of pH homeostasis may be of general importance for stress tolerance.

Understanding the genetic factors involved in seed longevity is of paramount importance in agricultural and ecological contexts. The polygenic nature of this trait suggests that many of them remain undiscovered. Here, we exploited the contrasting seed longevity found amongst Arabidopsis thaliana accessions to further understand this phenomenon. Concentrations of glutathione were higher in longer‐lived than shorter‐lived accessions, supporting that redox poise plays a prominent role in seed longevity. However, high seed permeability, normally associated with shorter longevity, is also present in long‐lived accessions. Dry seed transcriptome analysis indicated that the contribution to longevity of stored messenger RNA (mRNAs) is complex, including mainly accession‐specific mechanisms. The detrimental effect on longevity caused by other factors may be counterbalanced by higher levels of specific mRNAs stored in dry seeds, for instance those of heat‐shock proteins. Indeed, loss‐of‐function mutant analysis demonstrated that heat‐shock factors HSF1A and 1B contributed to longevity. Furthermore, mutants of the stress‐granule zinc‐finger protein TZF9 or the spliceosome subunits MOS4 or MAC3A/MAC3B, extended seed longevity, positioning RNA as a novel player in the regulation of seed viability. mRNAs of proteins with putative relevance to longevity were also abundant in shorter‐lived accessions, reinforcing the idea that resistance to ageing is determined by multiple factors.