The maintenance of cellular phosphate (Pi) homeostasis is of great importance in living organisms. The SPX domain-containing protein 1 (SPX1) proteins from both Arabidopsis and rice have been ...proposed to act as sensors of Pi status. The molecular signal indicating the cellular Pi status and regulating Pi homeostasis in plants, however, remains to be identified, as Pi itself does not bind to the SPX domain. Here, we report the identification of the inositol pyrophosphate InsP8 as a signaling molecule that regulates Pi homeostasis in Arabidopsis. Polyacrylamide gel electrophoresis profiling of InsPs revealed that InsP8 level positively correlates with cellular Pi concentration. We demonstrated that the homologs of diphosphoinositol pentakisphosphate kinase (PPIP5K), VIH1 and VIH2, function redundantly to synthesize InsP8, and that the vih1 vih2 double mutant overaccumulates Pi. SPX1 directly interacts with PHR1, the central regulator of Pi starvation responses, to inhibit its function under Pi-replete conditions. However, this interaction is compromised in the vih1 vih2 double mutant, resulting in the constitutive induction of Pi starvation-induced genes, indicating that plant cells cannot sense cellular Pi status without InsP8. Furthermore, we showed that InsP8 could directly bind to the SPX domain of SPX1 and is essential for the interaction between SPX1 and PHR1. Collectively, our study suggests that InsP8 is the intracellular Pi signaling molecule serving as the ligand of SPX1 for controlling Pi homeostasis in plants.
The SPX domain-containing protein SPX1 has been proposed as a Pi sensor; however, Pi itself does not bind to SPX1 as a ligand. The Pi signal in plants was hitherto unknown. This study reports that the inositol pyrophosphate InsP8 directly binds to SPX1and is essential for SPX1-mediated inhibition of PHR1, the central regulator of phosphate starvation responses, highlighting the signaling role of InsP8 in regulating Pi homeostasis in plants.
Plant vacuoles serve as the primary intracellular compartments for phosphorus (P) storage. The Oryza sativa genome contains three genes that encode SPX (
SYG1/
PHO81/
XPR1)‐MFS (
Major
Facility
...Superfamily) proteins (OsSPX‐MFS1‐3). The physiological roles of the three transporters under varying P conditions in laboratory and field are not known. To address this knowledge gap, we generated single, double and triple mutants for three OsSPX‐MFS genes. All the mutants except Osspx‐mfs2 display lower vacuolar Pi concentrations and OsSPX‐MFSs overexpression plant display higher Pi accumulation, demonstrating that all OsSPX‐MFSs are vacuolar Pi influx transporters. OsSPX‐MFS3 plays the dominant role based on the phenotypes of single mutants in terms of growth, vacuolar and tissue Pi concentrations. OsSPX‐MFS2 is the weakest and only functions as vacuole Pi sequestration in an Osspx‐mfs1/3 background. The vacuolar Pi sequestration capacity was severely impaired in Osspx‐mfs1/3 and Osspx‐mfs1/2/3, which resulted in increased Pi allocation to aerial organs. High P in the panicle impaired panicle and fertility in Osspx‐mfs1/3 and Osspx‐mfs1/2/3. Osspx‐mfs2 resulted in a more stable yield compared to the wild type under low P in field grown plants. The results suggest that alteration of vacuolar Pi sequestration may be a novel effective strategy to improve rice tolerance to low phosphorus in cropping systems.
To secure phosphorus (P) from soil, most land plants use a direct phosphate uptake pathway via root hairs and epidermis and an indirect phosphate uptake pathway via mycorrhizal symbiosis. The ...interaction between these two pathways is unclear. Here, we mapped a network between transcription factors and mycorrhizal symbiosis-related genes using Y1H. Intriguingly, this gene regulatory network is governed by the conserved P-sensing pathway, centered on phosphate starvation response (PHR) transcription factors. PHRs are required for mycorrhizal symbiosis and regulate symbiosis-related genes via the P1BS motif. SPX-domain proteins suppress OsPHR2-mediated induction of symbiosis-related genes and inhibit mycorrhizal infection. In contrast, plants overexpressing OsPHR2 show improved mycorrhizal infection and are partially resistant to P-mediated inhibition of symbiosis. Functional analyses of network nodes revealed co-regulation of hormonal signaling and mycorrhizal symbiosis. This network deciphers extensive regulation of mycorrhizal symbiosis by endogenous and exogenous signals and highlights co-option of the P-sensing pathway for mycorrhizal symbiosis.
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•A new map of the rice mycorrhizal symbiosis transcriptional regulatory network•PHR-centered network controls arbuscular mycorrhizal symbiosis•Phosphate regulates mycorrhizal symbiosis via the SPX-PHR-centered network•Co-option of the conserved P-sensing pathway for mycorrhizal symbiosis
An interconnected network of 266 transcription factors and promoters of 47 mycorrhiza-related genes highlights a PHR-centered arbuscular mycorrhizal symbiosis transcriptional regulatory network in rice, which connects the indirect mycorrhizal phosphate uptake pathway with the direct phosphate uptake pathway.
Phosphorus is an essential nutrient for plants. It is stored as inorganic phosphate (Pi) in the vacuoles of land plants but as inorganic polyphosphate (polyP) in chlorophyte algae. Although it is ...recognized that the SPX-Major Facilitator Superfamily (MFS) and VPE proteins are responsible for Pi influx and efflux, respectively, across the tonoplast in land plants, the mechanisms that underlie polyP homeostasis and the transition of phosphorus storage forms during the evolution of green plants remain unclear. In this study, we showed that CrPTC1, encoding a protein with both SPX and SLC (permease solute carrier 13) domains for Pi transport, and CrVTC4, encoding a protein with both SPX and vacuolar transporter chaperone (VTC) domains for polyP synthesis, are required for vacuolar polyP accumulation in the chlorophyte Chlamydomonas reinhardtii. Phylogenetic analysis showed that the SPX-SLC, SPX-VTC, and SPX-MFS proteins were present in the common ancestor of green plants (Viridiplantae). The SPX-SLC and SPX-VTC proteins are conserved among species that store phosphorus as vacuolar polyP and absent from genomes of plants that store phosphorus as vacuolar Pi. By contrast, SPX-MFS genes are present in the genomes of streptophytes that store phosphorus as Pi in the vacuoles. These results suggest that loss of SPX-SLC and SPX-VTC genes and functional conservation of SPX-MFS proteins during the evolution of streptophytes accompanied the change from ancestral polyP storage to Pi storage.
The change in vacuolar phosphorus storage form from polyP to Pi is an important transition during plant evolution. This study demonstrates that SPX-SLC and SPX-VTC proteins function in the vacuolar polyP homeostasis of chlorophytes. Further analyses show that their loss in streptophyte algae coincides with the transition from polyP to Pi, indicating that gene loss is important for the evolution of traits inherited by land plants.
Unlike most ancient microRNAs, which conservatively target homologous genes across species, microRNA827 (miR827) targets two different types of SPX (SYG1/PHO81/XPR1)-domain-containing genes, NITROGEN ...LIMITATION ADAPTATION (NLA) and PHOSPHATE TRANSPORTER 5 (PHT5), in Arabidopsis thaliana and Oryza sativa to regulate phosphate (Pi) transport and storage, respectively. However, how miR827 shifted its target preference and its evolutionary history are unknown.
Based on target prediction analysis, we found that in most angiosperms, miR827 conservatively targets PHT5 homologs, but in Brassicaceae and Cleomaceae it preferentially targets NLA homologs, and we provide evidence for the transition of target preference during Brassicales evolution. Intriguingly, we found a lineage-specific loss of the miR827-regulatory module in legumes.
Analysis of miR827-mediated cleavage efficiency and the expression of PHT5 in A. thaliana indicated that accumulation of mutations in the target site and the exclusion of the target site by alternative transcriptional initiation eliminated PHT5 targeting by miR827.
Here, we identified a transition of miR827 target preference during plant evolution and revealed the uniqueness of miR827-mediated regulation among conserved plant miRNAs. Despite the change in its target preference, upregulation of miR827 by Pi starvation and its role in regulating cellular Pi homeostasis were retained.
Phosphate, as a cellular energy currency, essentially drives most biochemical reactions defining living organisms, and thus its homeostasis must be tightly regulated. Investigation into the role of ...inositol pyrophosphates (PP-IPs) has provided a novel perspective on the regulation of phosphate homeostasis. Recent data suggest that metabolic and signaling interplay between PP-IPs, ATP, and inorganic polyphosphate (polyP) influences and is influenced by cellular phosphate homeostasis. Different studies have demonstrated that the SPX protein domain is a key component of proteins involved in phosphate metabolism. How PP-IPs control some aspects of phosphate homeostasis has become clearer with the recently acquired crystal structures of SPX domains. We review here recent studies on eukaryote phosphate homeostasis and provide insights into future research.
Many proteins involved in phosphate metabolism contain an SPX domain, raising the possibility that this domain may sense cellular phosphate status.
The recently resolved structure of several SPX domains revealed the presence of a basic binding region that recognizes PP-IPs. The binding of these molecules to the SPX domains of the VTC protein complex stimulated inorganic polyP synthesis.
Changes in phosphate availability alter ATP and PP-IP levels allowing them to transduce cellular phosphate status.
Phosphate homeostasis is also controlled by the ability of PP-IPs to regulate basic metabolism. The relative abundance, synthesis, and regulation of PP-IPs, ATP, and inorganic polyP are interconnected, affecting phosphate homeostasis and thus primary metabolism.
Phosphorus (P) is a crucial macronutrient for plant growth and development, and low-Pi stress poses a significant limitation to maize production. While the role of the SPX domain in encoding proteins ...involved in phosphate (Pi) homeostasis and signaling transduction has been extensively studied in other model plants, the molecular and functional characteristics of the SPX gene family members in maize remain largely unexplored. In this study, we identified six SPX members, and the phylogenetic analysis of ZmSPX s revealed a close relationship with SPX genes in rice. The promoter regions of ZmSPX s were abundant in biotic and abiotic stress-related elements, particularly associated with various hormone signaling pathways, indicating potential intersections between Pi signaling and hormone signaling pathways. Additionally, ZmSPX s displayed tissue-specific expression patterns, with significant and differential induction in anthers and roots, and were localized to the nucleus and cytoplasm. The interaction between ZmSPX s and ZmPHR s was established via yeast two-hybrid assays. Furthermore, overexpression of ZmSPX1 enhanced root sensitivity to Pi deficiency and high-Pi conditions in Arabidopsis thaliana . Phenotypic identification of the maize transgenic lines demonstrated the negative regulatory effect on the P concentration of stems and leaves as well as yield. Notably, polymorphic sites including 34 single-nucleotide polymorphisms (SNPs) and seven insertions/deletions (InDels) in ZmSPX1 were significantly associated with 16 traits of low-Pi tolerance index. Furthermore, significant sites were classified into five haplotypes, and haplotype5 can enhance biomass production by promoting root development. Taken together, our results suggested that ZmSPX family members possibly play a pivotal role in Pi stress signaling in plants by interacting with ZmPHR s. Significantly, ZmSPX1 was involved in the Pi-deficiency response verified in transgenic Arabidopsis and can affect the Pi concentration of maize tissues and yield. This work lays the groundwork for deeper exploration of the maize SPX family and could inform the development of maize varieties with improved Pi efficiency.
In plants, sensing the levels of external and internal nutrients is essential for reprogramming the transcriptome and adapting to the fluctuating environment. Phosphate (Pi) is a key plant nutrient, ...and a large proportion of Pi starvation-responsive genes are under the control of PHOSPHATE STARVATION RESPONSE REGULATOR 1 (PHR1) in Arabidopsis (AtPHR1) and its homologs, such as Oryza sativa (Os)PHR2 in rice. AtPHR1 and OsPHR2 expression is not very responsive to Pi starvation, raising the question as to how plants sense changes in cellular Pi levels to activate the central regulator. SPX named after SYG1 (suppressor of yeast gpa1 ), Pho81 (CDK inhibitor in yeast PHO pathway), and XPR1 (xenotropic and polytropic retrovirus receptor) proteins that harbor only the SPX domain are reported to be involved in the negative regulation of Pi starvation responses. Here, we show that the nuclear localized SPX proteins SPX1 and SPX2 are Pi-dependent inhibitors of the activity of OsPHR2 in rice. Indeed, SPX1 and SPX2 proteins interact with PHR2 through their SPX domain, inhibiting its binding to P1BS (the PHR1-binding sequence: GNATATNC). In vivo data, as well as results from in vitro experiments using purified SPX1, SPX2, and OsPHR2 proteins, showed that SPX1 and SPX2 inhibition of OsPHR2 activity is Pi-dependent. These data provide evidence to support the involvement of SPX1 and SPX2 in the Pi-sensing mechanism in plants.
Significance Phosphate (Pi) is a primary nutrient for plant growth. Because of the low availability of soil Pi, the Pi starvation signaling in plants is gaining great interest. Arabidopsis AtPHR1 and its rice homologue OsPHR2 are known to be central transcription factors in Pi homeostasis; however, the mechanism of how plants sense external Pi fluctuation to regulate the activity of AtPHR1/OsPHR2 has been elusive. Here, we identify rice SPX1 and SPX2 as Pi-dependent inhibitors of PHR2, implicating SPX1 and SPX2 in the Pi-sensing mechanism. We also show that the SPX domain of SPX1 and SPX2 is critical for repressing PHR2 binding to cis elements by protein interaction. The discovery of cellular nutrient concentration-dependent fine-tuning sheds light on a novel mechanism of plant adaption to environmental cues.
Arbuscular mycorrhizal fungi form symbiotic associations with most land plants and deliver mineral nutrients, in particular phosphate, to the host. Therefore, understanding the mechanisms of ...phosphate acquisition and delivery in the fungi is critical for full appreciation of the mutualism in this association. Here, we provide updates on physical, chemical, and biological strategies of the fungi for phosphate acquisition, including interactions with phosphate-solubilizing bacteria, and those on the regulatory mechanisms of phosphate homeostasis based on resurveys of published genome sequences and a transcriptome with reference to the latest findings in a model fungus. For the mechanisms underlying phosphate translocation and export to the host, which are major research frontiers in this field, not only recent advances but also testable hypotheses are proposed. Lastly, we briefly discuss applicability of the latest tools to gene silencing in the fungi, which will be breakthrough techniques for comprehensive understanding of the molecular basis of fungal phosphate metabolism.
Phosphorus (P) is an essential nutrient for plant growth and reproduction. Plants preferentially absorb P as orthophosphate (Pi), an ion that displays low solubility and that is readily fixed in the ...soil, making P limitation a condition common to many soils and Pi fertilization an inefficient practice. To cope with Pi limitation, plants have evolved a series of developmental and physiological responses, collectively known as the Pi starvation rescue system (PSR), aimed to improve Pi acquisition and use efficiency (PUE) and protect from Pi-starvation-induced stress. Intensive research has been carried out during the last 20 years to unravel the mechanisms underlying the control of the PSR in plants. Here we review the results of this research effort that have led to the identification and characterization of several core Pi starvation signaling components, including sensors, transcription factors, microRNAs (miRNAs) and miRNA inhibitors, kinases, phosphatases, and components of the proteostasis machinery. We also refer to recent results revealing the existence of intricate signaling interplays between Pi and other nutrients and antagonists, N, Fe, Zn, and As, that have changed the initial single-nutrient-centric view to a more integrated view of nutrient homeostasis. Finally, we discuss advances toward improving PUE and future research priorities.
Improving plant phosphorus acquisition and use efficiency (PUE) is required to implement a sustainable agriculture. This review discusses the plant signaling mechanisms controlling adaptation to low P nutrition and analyzes emerging data disclosing the integrated nature of nutrient homeostasis. The most recent advances toward PUE improvement and future research directions are also discussed.
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