RT Journal Article
SR Electronic
T1 Feedback inhibition of AMT1 NH<sub>4</sub><sup>+</sup>-transporters mediated by CIPK15 kinase
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2020.11.03.367383
DO 10.1101/2020.11.03.367383
A1 Chen, Hui-Yu
A1 Chen, Yen-Ning
A1 Wang, Hung-Yu
A1 Liu, Zong-Ta
A1 Frommer, Wolf B.
A1 Ho, Cheng-Hsun
YR 2020
UL http://biorxiv.org/content/early/2020/11/04/2020.11.03.367383.abstract
AB 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. 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−). 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.Competing Interest StatementThe authors have declared no competing interest.