PHO1: linking phosphate nutrition translocation and floral signalling in plants

This article comments on: Dai S, Chen H, Shi Y, Xiao X, Xu L, Qin C, Zhu Y, Yi K, Lei M, Zeng H. 2024. PHOSPHATE1-mediated phosphate translocation from roots to shoots regulates floral transition in plants. Journal of Experimental Botany 75, 5054–5075. https://doi.org/10.1093/jxb/erae222

Phosphorus is an essential macronutrient required for plant growth and development.A phosphate-responsive signalling pathway, known as the PHO pathway, has been extensively studied and many components were identified (Fig. 1).PHO1, encoding a SYG1/PHO81/XPR1-ERD1/XPR1/SYG1 (SPX-EXS) protein, is well known to be involved in loading of Pi into the root xylem vessels, and transferring Pi from roots to shoots in Arabidopsis (Hamburger et al., 2002).PHO1 protein abundance is adjusted via its degradation mediated by the ubiquitinconjugating E2 enzyme, PHOSPHATE2 (PHO2) (Liu et al., 2012).The PHO2 gene expression is post-transcriptionally repressed by phosphorus starvation-induced miRNA399 (miR399) with the induction of MYB transcription factors called PHOSPHATE STARVATION REGULATOR 1 (PHR1) and PHR1-like proteins (Bari et al., 2006).SPX1 and SPX2 are direct Pi-dependent inhibitors of PHR1 and negatively regulate its transcriptional activity in Arabidopsis (Puga et al., 2014).
The regulation of flowering time in plants involves the integration of environmental stimuli and internal signals (Kinoshita and Richter, 2020).Various genetic pathways and molecular components contribute to this intricate regulatory network (Kinoshita and Richter, 2020).The central regulator in the photoperiodic pathway is CONSTANS (CO), a B-box zinc-finger transcription factor, which induces the expression of FT.FT, in turn, acts as a mobile floral stimulus, moving from leaves to the shoot apex to initiate flower formation (Shim et al., 2017).The vernalization pathway involves the epigenetic down-regulation of FLOWERING LOCUS C (FLC) through exposure to prolonged cold temperature.FLC antagonizes the gibberellin (GA) pathway by repressing flowering promoters such as FT and SUPPRESSOR OF CONSTANS OVEREXPRESSION 1 (SOC1) (Sharma et al., 2020).
Nevertheless, the mechanism by which Pi nutrition is involved in floral transition regulation remains unclear.Using the pho1 mutants, the study demonstrates that phosphorus nutrition in plants is a key factor influencing flowering (Dai et al., 2024).Dai et al. (2024) found that two pho1 mutants exhibited delayed flowering compared with the wild type (WT) under both long-day and short-day conditions.The late flowering phenotype of pho1 mutants can be rescued by complementation with pPHO1:PHO1-GFP, which confirms that PHO1 plays a positive role in floral transition.By growing pho1 and WT plants hydroponically with varying Pi concentrations, or spraying KH 2 PO 4 solution onto the rosettes, the authors showed that the late flowering phenotype of pho1 mutants is associated with low shoot Pi concentration.Pi supplementation, particularly at shoot apices, partially rescued this phenotype.Low Pi treatments repressed the flowering time of WT plants, emphasizing the significant role of Pi availability in regulating flowering time (Dai et al., 2024).Using a reciprocal micrografting experiment with WT and pho1 mutant seedlings, the authors demonstrate that PHO1 mutation in the roots fully explains the late flowering phenotype of pho1 mutants.The molecular mechanisms underlying the modulation of flowering time by the upstream components of PHO1 were further investigated.The findings suggest a complex interplay of Pi signalling components in the regulation of flowering time (Dai et al., 2024).
An additional line of evidence for the link between Pi homeostasis and flowering time came from the identification of 15 genes associated with floral transition in the gene expression comparison of the pho1 mutant with the WT under KCl, or KH 2 PO 4 spray (Fig. 1).For instance, the expression of the well-known floral transition genes FT and TSF was dramatically reduced in pho1 compared with the WT, which further confirmed the link between flowering time and phosphorus nutrition.The late flowering phenotype of the pho1 mutant could be restored by overexpression of the FT gene.

Is jasmonate signalling involved in the interaction of phosphorus nutrition and flowering time?
It is well known that jasmonic acid (JA) homeostasis and signalling play diverse roles in plant reproduction (Yuan and Zhang, 2015).The biosynthetic and signalling pathways of JA are relatively conserved among different plant species, but the role of JA in reproductive development is (Yuan and Zhang, 2015;Han, 2017).In rice, JA promotes rice spikelet and stamen development.In contrast, JA signalling plays a negative role in floral transition in Arabidopsis (Zhao et al., 2022).Dai et al. (2024) found that many JA-responsive genes were differentially expressed in the pho1 mutant when sprayed with KH 2 PO 4 or KCl.This is consistent with the previous observation that Pi deficiency induces the JA pathway with involvement of PHO1 (Khan et al., 2016).The late flowering phenotype of pho1 mutants can be partially rescued by the mutation of CORONATINE INSENSITIVE 1 (COI1), which is a key component of the JA perception machinery, and partially recovered by the knockout of JAR1 (JA-RESISTANCE 1) that is involved in JA biosynthesis.Thus, the induction of the JA signalling pathway is partially responsible for the late flowering phenotype of pho1 mutants.Additionally, loss of function of OsPHO1;2, the rice homologue of PHO1, delays floral transition.The down-regulation of Hd3a and RFT1 (the rice homologues of FT) in the OsPHO1;2 mutant further underscores the conserved role of PHO1 homologues in regulating flowering time across plant species (Dai et al., 2024).

Future perspective
The plant hormone abscisic acid (ABA) and the balance of reactive oxygen species (ROS) in plant cells are associated with floral transition in plants (Wang et al., 2013;Huang et al., 2021).Gene Ontology analysis revealed enrichment of various biological processes and molecular functions among the differentially expressed genes in the pho1 mutant compared with the WT, indicating that the impact of PHO1 knockout might be not only on the JA pathway.The ABA-related genes AIK1 (ABA-insensitive protein kinase I) and AIRP2 (ABA-insensitive RING protein 2), the auxin biosynthesis genes YUC8 and YUC9, and the ROS homeostasis-related gene ARS1 (ABA & ROS sensitive 1) were found to be differentially expressed in pho1 compared with the WT (Dai et al., 2024).Thus, the loss of function of PHO1 or changes in external Pi supply could influence ABA, auxin, and ROS responses, which may in turn affect the floral transition.Whether and how these hormones and cellular processes could rescue the late flowering phenotype of the pho1 mutants remains to be determined in future studies.
Notably, Dai et al. (2024) found that knockout of PHO2, the upstream negative regulator of PHO1, led to early flowering under long-day conditions in Arabidopsis.However, rice pho2 was shown to have delayed flowering phenotypes (Li et al., 2017).As the mechanism of post-translational degradation of PHO2 on PHO1 is common between rice and Arabidopsis (Liu et al., 2012), how did the difference in the effect of PHO2 on flowering in the two species arise?Is this related to the different effects of JA on flowering, or is the effect of PHO2 on regulation of the circadian clock and photoperiod-responsive genes between the long-day plant Arabidopsis and the shortday plant rice?Analysis of the expression of JA-responsive and flowering-related genes as well as involvement of the photoperiod pathway in both rice and Arabidopsis pho2 mutants may solve the puzzle.
Single-cell and spatial transcriptomics are advanced techniques used in plant biology to understand gene expression at a highly detailed level (Rao et al., 2021).Spatial transcriptomics can provide insights into how PHO1-mediated Pi translocation regulates floral transition in plants on a cellular level, how Pi-responsive genes are expressed in different tissues, how neighbouring cells interact in response to different Pi levels, and how the overall tissue architecture adapts to changes in Pi availability and regulates floral transition.
Overall, this study significantly contributes to our understanding of the function of PHO1 and phosphate translocation in floral transition and provides valuable insights into the molecular mechanisms governing flowering time regulation in plants.The proposed model for how PHO1-mediated phosphate translocation and signalling regulates floral transition in plants and future perspective is summarized in Fig. 1.

Fig. 1 .
Fig. 1.Defining and dissecting PHO1-mediated floral transition pathways in plants.The integration of the phosphate signalling pathway with floral transition by the action of PHOSPHATE 1 (PHO1), the jasmonic acid (JA) signalling pathway, and flowering pathway activators, repressors, as well as integrators.SYG1/PHO81/XPR1 (SPX)1 and SPX2 have a cellular Pi-dependent inhibitory effect on PHR1, and the expression of PHO2 is posttranscriptionally repressed by miR399, and PHO2 regulates the stability of PHO1 by mediating its degradation.The expression of flowering pathway integrator FLOWERING LOCUS T (FT) is repressed in the pho1 mutant and there is potential involvement of CONSTANS (CO) in PHO1-mediated flowering regulation under long-day conditions.Another flowering pathway integrator SHORT VEGETATIVE PHASE (SVP) may be required for the late flowering of pho1 mutants.Five genes encoding activators of floral transition, namely TARGET OF FLC AND SVP1 (TFS1), LEAFY (LFY), FRUITFULL (FUL), SEPALLATA3 (SEP3), and APETALA1 (AP1), were attenuated and four genes encoding repressors of floral transition, namely SCHLAFMUTZE (SMZ), NUCLEAR TRANSCRIPTION FACTOR Y SUBUNIT A-4 (NFYA4), RGA-LIKE 2 (RGL2), and MYC4, were induced in pho1 shoots.The induction of the JA signalling pathway is responsible for the late flowering phenotype of pho1 mutants with the involvement of CORONATINE INSENSITIVE 1 (COI1)-dependent JA signalling pathway as well as JAR1 (Dai et al., 2024).Future research could target three aspects: (i) whether other hormones and cellular processes could be involved in PHO1-mediated phosphate translocation regulating floral transition; (ii) the similarities and differences of PHO2, PHO1, and phosphate homeostasis-related components among different plant species and crops involved in floral transition; and (iii) using spatial transcriptomics to provide insights into how PHO1-mediated phosphate translocation regulates the floral transition in plants on a cellular level.Activators of floral transition are highlighted in red, and repressors of floral transition are indicated in blue.Arrows represent activation processes, and lines ending in bars indicate suppression processes.Paths illustrated with solid lines represent cases where experimental evidence exists for involvement; the dotted line indicates hypothetical involvement.The figure was generated with BioRender (https://biorender.com).