The regulatory module MdBT2–MdMYB88/MdMYB124–MdNRTs regulates nitrogen usage in Apple

Less than 40% of the nitrogen (N) fertilizer applied to soil is absorbed by crops. Thus, improving the N use efficiency of crops is critical for agricultural development. However, the underlying regulation of these processes remains largely unknown, particularly in woody plants. By conducting yeast two-hybrid assays, we identified one interacting protein of MdMYB88 and MdMYB124 in apple (Malus domestica), namely BTB and TAZ domain protein 2 (MdBT2). Ubiquitination and protein stabilization analysis revealed that MdBT2 ubiquitinates and degrades MdMYB88 and MdMYB124 via the 26S proteasome pathway. MdBT2 negatively regulates nitrogen usage as revealed by the reduced fresh weight, dry weight, N concentration, and N usage index of MdBT2 overexpression calli under low-N conditions. In contrast, MdMYB88 and MdMYB124 increase nitrate absorption, allocation, and remobilization by regulating expression of MdNRT2.4, MdNRT1.8, MdNRT1.7, and MdNRT1.5 under N limitation, thereby regulating N usage. The results obtained illustrate the mechanism of a regulatory module comprising MdBT2–MdMYB88/MdMYB124–MdNRTs, through which plants modulate N usage. These data contribute to a molecular approach to improve the N usage of fruit crops under limited N acquisition.