Five Post-Translational Modification Residues of CmPT2 Play Key Roles in Yeast and Rice

Chrysanthemum (Chrysanthemum morifolium Ramat.) is one of the largest cut flowers in the world. Phosphate transporter Pht1 family member CmPht1;2 protein (CmPT2) plays an important role in response to low-phosphate (LP) stress in chrysanthemum. Post-translational modification (PTM) can modulate the function of proteins in multiple ways. Here, we used yeast and rice systems to study the role of putative PTM in CmPT2 by determining the effect of mutation of key amino acid residues of putative glycosylation, phosphorylation, and myristoylation sites. We chose nine amino acid residues in the putative PTM sites and mutated them to alanine (A) (Cmphts). CmPT2 recovered the growth of yeast strain MB192 under LP conditions. However, G84A, G222A, T239A, Y242A, and N422A mutants could not grow normally under LP conditions. Analysis of phosphorus absorption kinetics showed that the Km of CmPT2 was 65.7 μM. Among the nine Cmphts, the expression of five with larger Km (124.4-397.5 μM) than CmPT2 was further evaluated in rice. Overexpression of CmPT2-OE increased plant height, effective panicle numbers, branch numbers, and yield compared with that of wild type 'Wuyunjing No. 7' (W7). Overexpression of Cmphts-OE led to decreased plant height and effective panicle numbers compared with that of the CmPT2-OE strain. The Pi content in roots of CmPT2-OE was higher than that of the W7 under both high (normal) phosphate (HP) and LP conditions. However, the Pi content in the leaves and roots was significantly lower in the N422A-OE strain than in the CmPT2-OE strain under both HP and LP conditions. Under LP conditions, the phosphorus starvation response (PSR) genes in CmPT2-OE were inhibited at the transcription level. The expression patterns of phosphorus-related genes in T239A, Y242A, and N422A-OE under LP conditions were different from those of CmPT2-OE. In conclusion, these five post-translational modification residues of CmPT2 play key roles in modulating the function of CmPT2. This work boosters our understanding of the function of phosphate transporters and provides genetic resources for improving the efficiency of phosphorus utilization in crop plants.