Signaling pathways involved in the phosphorylation of membrane proteins in response to N and C

Nitrogen is an essential macronutrient for plant growth. In addition, nitrogen also has signaling functions by triggering specific adaptations in central metabolism and root development. While basic uptake mechanisms for nitrate through plasma membrane located transport systems are well known and global transcriptional responses to changes in nitrogen (N) and carbon (C) availability have been studied in the past, the detailed mechanisms in regulation of NO3- uptake, as well as signal transduction events upstream of transcription are still barely revealed. Our work will aim at closing this gap by particularly focusing on studying the regulation of nitrate transporter NRT2.1 and nitrate responsive protein kinases. 

Phosphorylation is the most well studied post-translational modification with regulatory function. It can induce changes in protein activity, provide docking sites for protein-protein interaction or induce changes in subcellular location. Thus, modification-dependent changes in protein abundance, activity and subsequent modification-dependent protein-protein interactions are important features in signalling networks. NO3- transporters were in the past shown to be subject to protein phosphorylation, and especially for NRT1.1 the role of protein phosphorylation in switching of transport affinity has been well described. Also for ammonium transporters, a phosphorylation-dependent activation/inactivation mechanism was elucidated in recent years. Therefore, a particular focus of this project is to study the role of protein phosphorylation for the regulation of the major nitrate transporter NRT2.1.

Therefore, the scientific aim of this collaborative project is (i) to characterize the condition-dependent phosphorylation of target proteins related to N metabolism in response to C and N availability. It will involve both the identification novel C/N regulated proteins as well as the specific characterisation of the role of the known NRT2.1 phosphorylation sites in the regulation of root NO3- uptake, (ii) finding candidate protein kinases and phosphatases using both NRT2.1 and selected N-metabolism related proteins as targets and (iii) characterising in planta the impact of the interesting protein kinases and phosphatases on the phosphorylation of NRT2.1 and selected N metabolism related proteins and the regulation of root NO3-uptake. We will also start to investigate the proteins interacting with the selected protein kinases and phosphatases as baits. We are convinced that with the complementary expertise of both partner groups and with a clear focus on NRT2.1 regulation at the protein level, we will gain novel valuable insights into a crucial plant function, namely the regulation of N uptake.