Tissue-specific respiratory burst oxidase homolog-dependent H2O2 signaling to the plasma membrane H+-ATPase confers potassium uptake and salinity tolerance in Cucurbitaceae

Abstract

Potassium (K+) is a critical determinant of salinity tolerance, and H2O2 has been recognized as an important signaling molecule that mediates many physiological responses. However, the details of how H2O2 signaling regulates K+ uptake in the root under salt stress remain elusive. In this study, salt-sensitive cucumber and salt-tolerant pumpkin which belong to the same family, Cucurbitaceae, were used to answer the above question. We show that higher salt tolerance in pumpkin was related to its superior ability for K+ uptake and higher H2O2 accumulation in the root apex. Transcriptome analysis showed that salinity induced 5816 (3005 up- and 2811 down-) and 4679 (3965 up- and 714 down-) differentially expressed genes (DEGs) in cucumber and pumpkin, respectively. DEGs encoding NADPH oxidase (respiratory burst oxidase homolog D; RBOHD), 14-3-3 protein (GRF12), plasma membrane H+-ATPase (AHA1), and potassium transporter (HAK5) showed higher expression in pumpkin than in cucumber under salinity stress. Treatment with the NADPH oxidase inhibitor diphenylene iodonium resulted in lower RBOHD, GRF12, AHA1, and HAK5 expression, reduced plasma membrane H+-ATPase activity, and lower K+ uptake, leading to a loss of the salinity tolerance trait in pumpkin. The opposite results were obtained when the plants were pre-treated with exogenous H2O2. Knocking out of RBOHD in pumpkin by CRISPR/Cas9 [clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9] editing of coding sequences resulted in lower root apex H2O2 and K+ content and GRF12, AHA1, and HAK5 expression, ultimately resulting in a salt-sensitive phenotype. However, ectopic expression of pumpkin RBOHD in Arabidopsis led to the opposite effect. Taken together, this study shows that RBOHD-dependent H2O2 signaling in the root apex is important for pumpkin salt tolerance and suggests a novel mechanism that confers this trait, namely RBOHD-mediated transcriptional and post-translational activation of plasma membrane H+-ATPase operating upstream of HAK5 K+ uptake transporters.

HvAKT2 and HvHAK1 confer drought tolerance in barley through enhanced leaf mesophyll H+ homoeostasis

“…Knocking out of RBOHD in pumpkin via genome editing led to a saltsensitive phenotype: lower root apex H 2 O 2 and K + content and GRF12, AHA1 and HAK5 expression. Therefore, salt tolerance in pumpkin is regulated by RBOHD-dependent transcriptional and post-translational activation of AHAs operating upstream of HAK5 (Huang et al, 2019). In this study, H 2 O 2 content increased significantly in plants silencing HvAKT2 and HvHAK1, but significantly reduced in overexpression plants under drought stress, which were also linked to altered K + and H + homoeostasis and drought tolerance in barley (Figures 4-8).…”

“…H 2 O 2 is an early response of K + deficiency and externally added H 2 O 2 was sufficient for the induction of the high-affinity K + uptake (Shin and Schachtman, 2004;Wang et al, 2017). Although there was no report on the correlation between AKT2 and HAK1 regulated K + homoeostasis and H 2 O 2 accumulation in plant drought tolerance, positive relationships between H 2 O 2 production and K + accumulation under salt stress were found for Arabidopsis, cucumber (Cucumis sativus) and pumpkin (Cucurbita moschata) (Huang et al, 2019;Ma et al, 2012;Redwan et al, 2016). For instance, it was demonstrated that higher salt tolerance in pumpkin is related to its higher K + uptake and H 2 O 2 accumulation in the root apex.…”

Tissue Tolerance Coupled With Ionic Discrimination Can Potentially Minimize the Energy Cost of Salinity Tolerance in Rice

“…Rapid induction of plasma membrane and vacuolar proton pumps to maintain negative membrane potential and to facilitate Na + /H + pumps was reported in salt-tolerant genotypes under stress (Vera-Estrella et al, 2005;Shabala and Mackay, 2011). Both Na + exclusion and K + uptake from rhizospheric region and selective upward transport mediated by xylem unloading is energy dependent and supported by increased action of ATPases and pyrophosphatases (Chakraborty et al, 2018;Niu et al, 2018a;Huang et al, 2019). To date, there are 11 variants of AHA (a plasma membrane ATPase) reported in rice (Ueno et al, 2005).…”

Nitric Oxide Enhances Cytotoxicity of Lead by Modulating the Generation of Reactive Oxygen Species and Is Involved in the Regulation of Pb2+ and Ca2+ Fluxes in Tobacco BY-2 Cells

“…Most reports indicate that exogenous NO supplementation has a role in the protection of plants by alleviating heavy metal stress, including Cd [50], Cu [51], Pb [52], and so forth. There are a few reports showing that the application of exogenous NO in combination with heavy metals enhances metal toxicity [53]. The contribution of endogenous NO to plant metal stress was also reported to exert both cytotoxic and cytoprotective effects [14].…”

“…NMT has been reported to be an effective approach to studying ion uptake and accumulation in plants and animals. The fluxes of ions, such as Cd 2+ , Ca 2+ , K + , Pb 2+ , and so forth, can be measured by NMT under normal physiological conditions [31,53,54]. Heavy metal transport is crucial for understanding metal uptake mechanisms in plants.…”