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

• Published in: BMC biology Vol. 18; no. 1; pp. 1 - 13
• Main Authors: Chen, Hui-Yu, Chen, Yen-Ning, Wang, Hung-Yu, Liu, Zong-Ta, Frommer, Wolf B., Ho, Cheng-Hsun
• Format: Journal Article
• Language: English
• Published: London BioMed Central 14.12.2020; BioMed Central Ltd; BMC
• Subjects: Allosteric properties; Ammonia; Ammonium; Analysis; Arabidopsis thaliana; Bacteria; Biomedical and Life Sciences; Biosensors; C-Terminus; Carrier proteins; Chemical properties; Control; Feedback control; Feedback inhibition; Gametocytes; Influence; Kinases; Life Sciences; Nitrogen; Nutrition; Oocytes; Phosphorylation; Protein kinase; Protein kinase C; Protein kinases; Proteins; Research Article; Seedlings; Sensors; Threonine; Transcription; Transporter; Yeast; Taiwan; Ammonium; Phosphorylation; Protein kinase; Transporter
• ISSN: 1741-7007; 1741-7007
• DOI: 10.1186/s12915-020-00934-w

Background Ammonium (NH 4 + ), a key nitrogen form, becomes toxic when it accumulates to high levels. Ammonium transporters (AMTs) are the key transporters responsible for NH 4 + 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 NH 4 + -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 NH 4 + 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 NH 4 + levels. Notably, cipk15 knockout mutants showed higher 15 NH 4 + uptake and accumulated higher amounts of NH 4 + compared to the wild-type. Consistently, cipk15 was hypersensitive to both NH 4 + and methylammonium but not nitrate (NO 3 − ). 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 NH 4 + -accumulation.