Crystal structure of the plant dual-affinity nitrate transporter NRT1.1

Nitrate is a primary nutrient for plant growth, but its levels in soil can fluctuate by several orders of magnitude. Previous studies have identified Arabidopsis NRT1.1 as a dual-affinity nitrate transporter that can take up nitrate over a wide range of concentrations. The mode of action of NRT1.1 is controlled by phosphorylation of a key residue, Thr 101; however, how this post-translational modification switches the transporter between two affinity states remains unclear. Here we report the crystal structure of unphosphorylated NRT1.1, which reveals an unexpected homodimer in the inward-facing conformation. In this low-affinity state, the Thr 101 phosphorylation site is embedded in a pocket immediately adjacent to the dimer interface, linking the phosphorylation status of the transporter to its oligomeric state. Using a cell-based fluorescence resonance energy transfer assay, we show that functional NRT1.1 dimerizes in the cell membrane and that the phosphomimetic mutation of Thr 101 converts the protein into a monophasic high-affinity transporter by structurally decoupling the dimer. Together with analyses of the substrate transport tunnel, our results establish a phosphorylation-controlled dimerization switch that allows NRT1.1 to uptake nitrate with two distinct affinity modes. A description of the crystal structure of unphosphorylated NRT1.1 provides insights into how phosphorylation switches the nitrate transporter between the low-affinity and high-affinity states. Soil levels of nitrate, a primary nutrient for plant growth, can vary dramatically. Plants therefore need a versatile mechanism for obtaining nitrate from the environment. In the model plant Arabidopsis thaliana, the dual-affinity transporter NRT1.1 can take up nitrate across a broad range of concentrations, switching from low- to high-affinity mode according to the phosphorylation status of a key threonine residue. Two studies published in this issue of Nature describe the crystal structures of full-length NRT1.1, providing insights into how this post-translational modification switches the transporter between the low-affinity and high-affinity states.