Transceptor NRT1.1 and receptor-kinase QSK1 complex controls PM H+-ATPase activity under low nitrate.

NRT1.1, a nitrate transceptor, plays an important role in nitrate binding, sensing, and nitrate-dependent lateral root (LR) morphology. However, little is known about NRT1.1-mediated nitrate signaling transduction through plasma membrane (PM)-localized proteins. Through in-depth phosphoproteome profiling using membranes of Arabidopsis roots, we identified receptor kinase QSK1 and plasma membrane H+-ATPase AHA2 as potential downstream components of NRT1.1 signaling in a mild low-nitrate (LN)-dependent manner. QSK1, as a functional kinase and molecular link, physically interacts with NRT1.1 and AHA2 at LN and specifically phosphorylates AHA2 at S899. Importantly, we found that LN, not high nitrate (HN), induces formation of the NRT1.1-QSK1-AHA2 complex in order to repress the proton efflux into the apoplast by increased phosphorylation of AHA2 at S899. Loss of either NRT1.1 or QSK1 thus results in a higher T947/S899 phosphorylation ratio on AHA2, leading to enhanced pump activity and longer LRs under LN. Our results uncover a regulatory mechanism in which NRT1.1, under LN conditions, promotes coreceptor QSK1 phosphorylation and enhances the NRT1.1-QSK1 complex formation to transduce LN sensing to the PM H+-ATPase AHA2, controlling the phosphorylation ratio of activating and inhibitory phosphorylation sites on AHA2. This then results in altered proton pump activity, apoplast acidification, and regulation of NRT1.1-mediated LR growth.

Protein Dynamics in Young Maize Root Hairs in Response to Macro- and Micronutrient Deprivation.

Plants increase their root surface with root hairs to improve the acquisition of nutrients from the soil. The unicellular character of root hairs and their position at the root surface make them an attractive system to investigate adaptive processes of rhizodermal cells that are in direct contact with the soil solution. In young maize seedlings, roots are densely covered with root hairs, although nutrient reserves in the seed are sufficient to support seedling growth rates for a few days. We used a label-free quantitative proteomics approach to study protein abundance adjustments in 4 day old root hairs grown in aeroponic culture in the presence and absence of several macro- and micronutrients. Compared to the proteome of root hairs developed under full nutrition, protein abundance changes were observed in pathways related to macronutrient (N, P, K, and Mg) deficiencies. For example, lack of N in the medium repressed the primary N metabolism pathway, increased amino acid synthesis, but repressed their degradation, and affected the primary carbon metabolism, such as glycolysis. Glycolysis was similarly affected by K and P deprivation, but the glycolytic pathway was negatively regulated by the absence of the micronutrients Fe and Zn. In contrast, the deprivation of Mn had almost no affect on the root hair proteome. Our results indicate either that the metabolism of very young root hairs adjusts to cellular nutrient deficiencies that have been already experienced or that root hairs sense the external lack of specific nutrients in the nutrient solution and adjust their metabolism accordingly.

Role of the Npr1 kinase in ammonium transport and signaling by the ammonium permease Mep2 in Candida albicans.

The ammonium permease Mep2 induces a switch from unicellular yeast to filamentous growth in response to nitrogen limitation in Saccharomyces cerevisiae and Candida albicans. In S. cerevisiae, the function of Mep2 and other ammonium permeases depends on the protein kinase Npr1. Mutants lacking NPR1 cannot grow on low concentrations of ammonium and do not filament under limiting nitrogen conditions. A G349C mutation in Mep2 renders the protein independent of Npr1 and results in increased ammonium transport and hyperfilamentous growth, suggesting that the signaling activity of Mep2 directly correlates with its ammonium transport activity. In this study, we investigated the role of Npr1 in ammonium transport and Mep2-mediated filamentation in C. albicans. We found that the two ammonium permeases Mep1 and Mep2 of C. albicans differ in their dependency on Npr1. While Mep1 could function well in the absence of the Npr1 kinase, ammonium transport by Mep2 was virtually abolished in npr1Δ mutants. However, the dependence of Mep2 activity on Npr1 was relieved at higher temperatures (37°C), and Mep2 could efficiently induce filamentous growth under limiting nitrogen conditions in npr1Δ mutants. Like in S. cerevisiae, mutation of the conserved glycine at position 343 in Mep2 of C. albicans to cysteine resulted in Npr1-independent ammonium uptake. In striking contrast, however, the mutation abolished the ability of Mep2 to induce filamentous growth both in the wild type and in npr1Δ mutants. Therefore, a mutation that improves ammonium transport by Mep2 under nonpermissible conditions eliminates its signaling activity in C. albicans.