Dissecting the Roles of MYB-related Transcription Factor PRF1 and High-Affinity Pi Transporter Pht1;5 in Pathways Regulating Phosphate Mobilization in Arabidopsis

Phosphate (Pi) is an integral component of numerous biological molecules and a constituent of major metabolic processes during plant growth and development. Pi deficiency is one of the key factors limiting crop productivity. A thorough analysis of the molecular mechanisms of Pi acquisition and mobilization would help us understand how plants adapt to low Pi levels. Several molecular determinants including transcription factors (TFs) have been shown to regulate Pi transport and other Pi-starvation adaptive responses. This study is focused on functional analysis of a MYB-related GARP TF, PRF1 (Pi-STARVATION REGULATING FACTOR 1) and its interaction with PHR1 (PHOSPHATE STARVATION RESPONSE 1), a key TF regulating Pi signaling in Arabidopsis. PRF1 was identified from a Pi-responsive microarray study and it was found to be induced in the Pi-deprived roots and shoots of Arabidopsis. Further, its nuclear localization was confirmed by GFP::PRF1 in transgenic plants. Loss- of-function and overexpression of PRF1 revealed that it positively regulated a subset of high-affinity Pi transporters and members of the PHR1-regulon that control Pi distribution. Additionally PRF1 also appears to participate in root development independent of the Pi regime. The double mutant phr1/prf1 was identical to phr1 with respect to shoot Pi content, anthocyanin accumulation and expression of members of PHR1-regulon. This suggested that PRF1 functions downstream of PHR1 in the Pi signaling pathway and positively regulates Pi transport and mobilization. One of the high-affinity Pi transporters, Pht1;5 was strongly induced in the roots of 35S::PRF1 under Pi-replete conditions, suggesting that it could be responsible for the altered Pi mobilization between root/shoot observed for these plants. Distribution of internal Pi is crucial for sustaining growth of new leaves and development of seeds. Pi-starvation induced Pht1;5 is expressed in the phloem cells of senescing leaves and thought to play a role in Pi remobilization from older leaves to metabolically active tissues of the plant. Kinetic analysis using 33 Pi uptake and P content of the seedlings with either loss-of-function or constitutive overexpression of Pht1;5 seedlings revealed that this gene is involved in Pi transfer from root to shoot but not in acquisition of Pi from the rhizosphere. Plants overexpressing Pht1;5 also showed premature senescence compared to the wild-type in addition to increased Pi remobilization out of older leaves. Further, Pht1;5-overexpressors showed enhanced root-hair formation as well as reduced primary root that could be rescued by the application of ethylene production inhibitor, aminoethoxyvinyl-glycine (AVG). Taken together these results suggest a role for Pht1;5 in Pi translocation and remobilization within the plant. This also establishes a tangible connection between senescence, ethylene and the regulation of Pi transporters. Thus this work sheds light on the intricate molecular and genetic mechanisms regulating Pi distribution within the plant