CPK28-mediated phosphorylation enhances nitrate transport activity of NRT2.1 during nitrogen deprivation.

Autor: Yue L; State Key Laboratory of Plant Environmental Resilience, College of Biological Sciences, China Agricultural University, Beijing, 100193, China., Liu M; Beijing Key Laboratory of Maize DNA Fingerprinting and Molecular Breeding, Beijing Academy of Agriculture & Forestry Sciences, Beijing, 100097, China., Liao J; State Key Laboratory of Plant Environmental Resilience, College of Biological Sciences, China Agricultural University, Beijing, 100193, China., Zhang K; State Key Laboratory of Plant Environmental Resilience, College of Biological Sciences, China Agricultural University, Beijing, 100193, China., Wu WH; State Key Laboratory of Plant Environmental Resilience, College of Biological Sciences, China Agricultural University, Beijing, 100193, China., Wang Y; State Key Laboratory of Plant Environmental Resilience, College of Biological Sciences, China Agricultural University, Beijing, 100193, China.
Jazyk: angličtina
Zdroj: The New phytologist [New Phytol] 2025 Jan; Vol. 245 (1), pp. 249-262. Date of Electronic Publication: 2024 Nov 02.
DOI: 10.1111/nph.20236
Abstrakt: Nitrate (NO 3 - ) serves as the primary inorganic nitrogen source assimilated by most terrestrial plants. The acquisition of nitrate from the soil is facilitated by NITRATE TRANSPORTERS (NRTs), with NRT2.1 being the key high-affinity nitrate transporter. The activity of NRT2.1, which has multiple potential phosphorylation sites, is intricately regulated under various physiological conditions. Here, we discovered that CALCIUM-DEPENDENT PROTEIN KINASE 28 (CPK28) positively regulates nitrate uptake under nitrogen deprivation conditions. We found CPK28 as the kinase targeted by immunoprecipitation followed by mass spectrometry and examined the in-planta phosphorylation status of NRT2.1 in cpk28 mutant plants by employing quantitative MS-based phosphoproteomics. Through a combination of in vitro phosphorylation experiment and immunoblotting using phospho-specific antibody, we successfully demonstrated that CPK28 specifically phosphorylates NRT2.1 at Ser21. Functional analysis conducted in Xenopus oocytes revealed that co-expression of CPK28 significantly enhanced high-affinity nitrate uptake of NRT2.1. Further investigation using transgenic plants showed that the phosphomimic variant NRT2.1 S21E , but not the nonphosphorylatable variant NRT2.1 S21A , fully restored high-affinity 15 NO 3 - uptake ability in both nrt2.1 and cpk28 mutant backgrounds. This study clarifies that the kinase activity of CPK28 is promoted during nitrogen deprivation conditions. These significant findings provide valuable insights into the intricate regulatory mechanisms that govern nitrate-demand adaptation.
(© 2024 The Author(s). New Phytologist © 2024 New Phytologist Foundation.)
Databáze: MEDLINE
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