MicroRNA399 is a long-distance signal for the regulation of plant phosphate homeostasis

The presence of microRNA species in plant phloem sap suggests potential signaling roles by long-distance regulation of gene expression. Proof for such a role for a phloem-mobile microRNA is lacking. Here we show that phosphate (Pi) starvation-induced microRNA399 (miR399) is present in the phloem sap of two diverse plant species, rapeseed and pumpkin, and levels are strongly and specifically increased in phloem sap during Pi deprivation. By performing micro-grafting experiments using Arabidopsis, we further show that chimeric plants constitutively over-expressing miR399 in the shoot accumulate mature miR399 species to very high levels in their wild-type roots, while corresponding primary transcripts are virtually absent in roots, demonstrating shoot-to-root transport. The chimeric plants exhibit (i) down-regulation of the miR399 target transcript (PHO2), which encodes a critical component for maintenance of Pi homeostasis, in the wild-type root, and (ii) Pi accumulation in the shoot, which is the phenotype of pho2 mutants, miR399 over-expressers or chimeric plants with a genetic knock-out of PHO2 in the root. Hence the transported miR399 molecules retain biological activity. This is a demonstration of systemic control of a biological process, i.e. maintenance of plant Pi homeostasis, by a phloem-mobile microRNA

Phosphate acquisition and metabolism in plants.

Plants need at least 13 different nutrients to maintain optimal growth. Nitrogen and phosphorus, from the Greek 'phôs' (meaning 'light') and 'phoros' (meaning 'bearer'), are the main nutrients limiting plant growth in both agricultural and natural ecosystems. Agriculture has relied heavily since the mid 1950s on the use of synthetic ammonium- and phosphorus-based fertilizers to increase crop productivity. While industrial synthesis of ammonium relies on the chemical conversion of atmospheric nitrogen, phosphorus is mined from finite reserves concentrated in a few countries. Considering our current dependence on phosphorus fertilizers for food production and the geopolitical aspects associated with current resources, it will be important to develop technologies enabling the maintenance of high crop yield with reduced fertilizer input. This will require an in-depth knowledge on the various pathways that enable plants to acquire phosphorus from the soil and maximize its economical use for growth and reproduction. In this primer, we give an overview of the factors limiting phosphorus acquisition by plants and highlight various pathways and strategies plants have evolved at the level of development, metabolism and signal transduction to adapt to phosphorus deficiency

PHO1 Exports Phosphate from the Chalazal Seed Coat to the Embryo in Developing Arabidopsis Seeds.

Seed production requires the transfer of nutrients from the maternal seed coat to the filial endosperm and embryo. Because seed coat and filial tissues are symplasmically isolated, nutrients arriving in the seed coat via the phloem must be exported to the apoplast before reaching the embryo. Proteins implicated in the transfer of inorganic phosphate (Pi) from the seed coat to the embryo are unknown despite seed P content being an important agronomic trait. Here we show that the Arabidopsis Pi exporters PHO1 and PHOH1 are expressed in the chalazal seed coat (CZSC) of developing seeds. PHO1 is additionally expressed in developing ovules. Phosphorus (P) content and Pi flux between the seed coat and embryo were analyzed in seeds from grafts between WT roots and scions from either pho1, phoh1, or the pho1 phoh1 double mutant. Whereas P content and distribution between the seed coat and embryo in fully mature dry seeds of these mutants are similar to the WT, at the mature green stage of seed development the seed coat of the pho1 and pho1 phoh1 mutants, but not of the phoh1 mutant, retains approximately 2-fold more P than its WT control. Expression of PHO1 under a CZSC-specific promoter complemented the seed P distribution phenotype of the pho1 phoh1 double mutant. CZSC-specific down-expression of PHO1 also recapitulated the seed P distribution phenotype of pho1. Together, these experiments show that PHO1 expression in the CZSC is important for the transfer of P from the seed coat to the embryo in developing seeds

Modulation of Shoot Phosphate Level and Growth by PHOSPHATE1 Upstream Open Reading Frame.

Inorganic orthophosphate (Pi) is an essential nutrient for plant growth, and its availability strongly impacts crop yield. PHOSPHATE1 (PHO1) transfers Pi from root to shoot via Pi export into root xylem vessels. In this work, we demonstrate that an upstream open reading frame (uORF) present in the 5' untranslated region of the Arabidopsis (Arabidopsis thaliana) PHO1 inhibits its translation and influences Pi homeostasis. The presence of the uORF strongly inhibited the translation of a PHO1 5'UTR-luciferase construct in protoplasts. A point mutation removing the PHO1 uORF (ΔuORF) in transgenic Arabidopsis resulted in increased association of its mRNA with polysomes and led to higher PHO1 protein levels, independent of Pi availability. Interestingly, deletion of the uORF led to higher shoot Pi content and was associated with improved shoot growth under low external Pi supply and no deleterious effects under Pi-sufficient conditions. We further show that natural accessions lacking the PHO1 uORF exhibit higher PHO1 protein levels and shoot Pi content. Increased shoot Pi content was linked to the absence of the PHO1 uORF in a population of F2 segregants. We identified the PHO1 uORF in genomes of crops such as rice (Oryza sativa), maize (Zea mays), barley (Hordeum vulgare), and wheat (Triticum aesativum), and we verified the inhibitory effect of the rice PHO1 uORF on translation in protoplasts. Our work suggests that regulation of PHO1 expression via its uORF might be a genetic resource useful-both in natural populations and in the context of genome editing-toward improving plant growth under Pi-deficient conditions

Biallelic transcription of Igf2 and H19 in individual cells suggests a post-transcriptional contribution to genomic imprinting

AbstractThe H19 and insulin-like growth factor 2 (Igf2) genes in the mouse are models for genomic imprinting during development. The genes are located only 90 kb apart in the same transcriptional orientation [1], but are reciprocally imprinted: Igf2 is paternally expressed while H19 is maternally expressed. It has been suggested that expression of H19 and repression of Igf2 (or the converse) on a given chromosome are mechanistically linked and that the parental imprint operates at the level of transcription [2]. Although expression of Igf2 and H19 is thought to be monoallelic, the data have so far been obtained exclusively by looking at steady-state RNA levels using techniques that reflect the average activity of the genes in a cell population [3,4]. Here, we have adapted a fluorescent in situ hybridisation (FISH) method to detect nascent RNA molecules of Igf2 and H19 at the initial transcription sites in the nuclei of wild-type mouse embryonic liver cells. Nine different transcription patterns were observed, reflecting a high heterogeneity of transcription at the single-cell level. Our observations suggest that regulation of Igf2 and H19 by parental imprinting is much more complex than previously proposed and acts at both transcriptional and post-transcriptional levels

Mott Transition, Compressibility Divergence and P-T Phase Diagram of Layered Organic Superconductors: An Ultrasonic Investigation

The phase diagram of the organic superconductor $\kappa$-(BEDT-TTF)$_2$Cu[N(CN)$_2$Cl has been investigated by ultrasonic velocity measurements under helium gas pressure. Different phase transitions were identified trough several elastic anomalies characterized from isobaric and isothermal sweeps. Our data reveal two crossover lines that end on the critical point terminating the first-order Mott transition line. When the critical point is approached along these lines, we observe a dramatic softening of the velocity which is consistent with a diverging compressibility of the electronic degrees of freedom.Comment: 4 pages, 5 figure

Functional expression of PHO1 to the Golgi and trans-Golgi network and its role in export of inorganic phosphate.

Arabidopsis thaliana PHO1 is primarily expressed in the root vascular cylinder and is involved in the transfer of inorganic phosphate (Pi) from roots to shoots. To analyze the role of PHO1 in transport of Pi, we have generated transgenic plants expressing PHO1 in ectopic A. thaliana tissues using an estradiol-inducible promoter. Leaves treated with estradiol showed strong PHO1 expression, leading to detectable accumulation of PHO1 protein. Estradiol-mediated induction of PHO1 in leaves from soil-grown plants, in leaves and roots of plants grown in liquid culture, or in leaf mesophyll protoplasts, was all accompanied by the specific release of Pi to the extracellular medium as early as 2-3 h after addition of estradiol. Net Pi export triggered by PHO1 induction was enhanced by high extracellular Pi and weakly inhibited by the proton-ionophore carbonyl cyanide m-chlorophenylhydrazone. Expression of a PHO1-GFP construct complementing the pho1 mutant revealed GFP expression in punctate structures in the pericycle cells but no fluorescence at the plasma membrane. When expressed in onion epidermal cells or in tobacco mesophyll cells, PHO1-GFP was associated with similar punctate structures that co-localized with the Golgi/trans-Golgi network and uncharacterized vesicles. However, PHO1-GFP could be partially relocated to the plasma membrane in leaves infiltrated with a high-phosphate solution. Together, these results show that PHO1 can trigger Pi export in ectopic plant cells, strongly indicating that PHO1 is itself a Pi exporter. Interestingly, PHO1-mediated Pi export was associated with its localization to the Golgi and trans-Golgi networks, revealing a role for these organelles in Pi transport

Divergence of the Chaotic Layer Width and Strong Acceleration of the Spatial Chaotic Transport in Periodic Systems Driven by an Adiabatic ac Force

We show for the first time that a {\it weak} perturbation in a Hamiltonian system may lead to an arbitrarily {\it wide} chaotic layer and {\it fast} chaotic transport. This {\it generic} effect occurs in any spatially periodic Hamiltonian system subject to a sufficiently slow ac force. We explain it and develop an explicit theory for the layer width, verified in simulations. Chaotic spatial transport as well as applications to the diffusion of particles on surfaces, threshold devices and others are discussed.Comment: 4 pages including 3 EPS figures, this is an improved version of the paper (accepted to PRL, 2005

Coordination between zinc and phosphate homeostasis involves the transcription factor PHR1, the phosphate exporter PHO1, and its homologue PHO1;H3 in Arabidopsis.

Interactions between zinc (Zn) and phosphate (Pi) nutrition in plants have long been recognized, but little information is available on their molecular bases and biological significance. This work aimed at examining the effects of Zn deficiency on Pi accumulation in Arabidopsis thaliana and uncovering genes involved in the Zn-Pi synergy. Wild-type plants as well as mutants affected in Pi signalling and transport genes, namely the transcription factor PHR1, the E2-conjugase PHO2, and the Pi exporter PHO1, were examined. Zn deficiency caused an increase in shoot Pi content in the wild type as well as in the pho2 mutant, but not in the phr1 or pho1 mutants. This indicated that PHR1 and PHO1 participate in the coregulation of Zn and Pi homeostasis. Zn deprivation had a very limited effect on transcript levels of Pi-starvation-responsive genes such as AT4, IPS1, and microRNA399, or on of members of the high-affinity Pi transporter family PHT1. Interestingly, one of the PHO1 homologues, PHO1;H3, was upregulated in response to Zn deficiency. The expression pattern of PHO1 and PHO1;H3 were similar, both being expressed in cells of the root vascular cylinder and both localized to the Golgi when expressed transiently in tobacco cells. When grown in Zn-free medium, pho1;h3 mutant plants displayed higher Pi contents in the shoots than wild-type plants. This was, however, not observed in a pho1 pho1;h3 double mutant, suggesting that PHO1;H3 restricts root-to-shoot Pi transfer requiring PHO1 function for Pi homeostasis in response to Zn deficiency