Dataset Information

Feedback inhibition of AMT1 NH4+-transporters mediated by CIPK15 kinase

ABSTRACT:

Background

Ammonium (NH4+), a key nitrogen form, becomes toxic when it accumulates to high levels. Ammonium transporters (AMTs) are the key transporters responsible for NH4+ uptake. AMT activity is under allosteric feedback control, mediated by phosphorylation of a threonine in the cytosolic C-terminus (CCT). However, the kinases responsible for the NH4+-triggered phosphorylation remain unknown.

Results

In this study, a functional screen identified protein kinase CBL-Interacting Protein Kinase15 (CIPK15) as a negative regulator of AMT1;1 activity. CIPK15 was able to interact with several AMT1 paralogs at the plasma membrane. Analysis of AmTryoshka, an NH4+ transporter activity sensor for AMT1;3 in yeast, and a two-electrode-voltage-clamp (TEVC) of AMT1;1 in Xenopus oocytes showed that CIPK15 inhibits AMT activity. CIPK15 transcript levels increased when seedlings were exposed to elevated NH4+ levels. Notably, cipk15 knockout mutants showed higher 15NH4+ uptake and accumulated higher amounts of NH4+ compared to the wild-type. Consistently, cipk15 was hypersensitive to both NH4+ and methylammonium but not nitrate (NO3?).

Conclusion

Taken together, our data indicate that feedback inhibition of AMT1 activity is mediated by the protein kinase CIPK15 via phosphorylation of residues in the CCT to reduce NH4+-accumulation.

Supplementary information

The online version contains supplementary material available at 10.1186/s12915-020-00934-w.

SUBMITTER: Chen H

PROVIDER: S-EPMC7737296 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BackgroundIn northwest of China, mini Chinese cabbage (Brassica pekinensis) is highly valued by consumers, and is widely cultivated during winter in solar-greenhouses where low light (LL) fluence (between 85 and 150 μmol m-2 s-1 in day) is a major abiotic stress factor limiting plant growth and crop productivity. The mechanisms with which various NH4+: NO3- ratios affected growth and photosynthesis of mini Chinese cabbage under normal (200 μmol m-2 s-1) and low (100 μmol m-2 s-1) light conditions was investigated. The four solutions with different ratios of NH4+: NO3- applied were 0:100, 10:90, 15:85 and 25:75 with the set up in a glasshouse in hydroponic culture. The most appropriate NH4+: NO3- ratio that improved the tolerance of mini Chinese cabbage seedlings to LL was found in our current study.ResultsUnder low light, the application of NH4+: NO3- (10:90) significantly stimulated growth compared to only NO3- by increasing leaf area, canopy spread, biomass accumulation, and net photosynthetic rate. The increase in net photosynthetic rate was associated with an increase in: 1) maximum and effective quantum yield of PSII; 2) activities of Calvin cycle enzymes; and 3) levels of mRNA relative expression of several genes involved in Calvin cycle. In addition, glucose, fructose, sucrose, starch and total carbohydrate, which are the products of CO2 assimilation, accumulated most in the cabbage leaves that were supplied with NH4+: NO3- (10:90) under LL condition. Low light reduced the carbohydrate: nitrogen (C: N) ratio while the application of NH4+: NO3- (10:90) alleviated the negative effect of LL on C: N ratio mainly by increasing total carbohydrate contents.ConclusionsThe application of NH4+:NO3- (10:90) increased rbcL, rbcS, FBA, FBPase and TK expression and/or activities, enhanced photosynthesis, carbohydrate accumulation and improved the tolerance of mini Chinese cabbage seedlings to LL. The results of this study would provide theoretical basis and technical guidance for mini Chinese cabbage production. In practical production, the ratio of NH4+:NO3- should be adjusted with respect to light fluence for successful growing of mini Chinese cabbage.