Biological Analysis and Response to Low Phosphate Stress of Phosphate Transporter Family 1 (PHT1) Genes in Solanum tuberosum L.

Inorganic phosphate(Pi)is one of the main nutrients necessary for plant growth.Phosphate transporters mediate the acquisition,transport and recycling of phosphate,which is essential for plant growth and development.Although PHT1 has been reported in many plants at home and abroad,it is rarely studied in potato.Therefore,it is of great significance to study the PHT1 family members in order to understand the molecular response mechanism of potato in low phosphate state.In this study,a total of 6 potato PHT1 genes were identified and isolated.It was found that after the expression of different members of potato PHT1 gene,there were certain differences in amino acids and proteins,and the transmembrane domains ranged from 6 to 12.The difference in the secondary and tertiary protein structure of potato PHT1 also led to a difference in protein morphology.In addition,the expression of the PHT1 gene in potato increased obviously during 3~9 h of Pi deficiency stress.Overall,the expression levels of different genes in roots,stems and leaves are distinctly different,but the expression levels of the StPHT1;6 and StPHT1;10 genes are very high in roots,stems and leaves,indicating that these two genes may participate in the absorption of Pi in potato and play a role in Pi translocation.These two genes play a major role in the regulation of expression under short-term Pi deficiency stress.Our results provide an important reference for further understanding the evolution and function of potato phosphate transporters,and have important significance for improving the ability of potato to tolerate low Pi.

A mitochondrial phosphate transporter,McPht gene,confers an acclimation regulation of the transgenic rice to phosphorus deficiency

Phosphate transporters play an important role in promoting the uptake and transport of phosphate in plants. In this study, the McPht gene from the Mesembryanthemum crystallinum, a mitochondrial phosphate transporter, was isolated and constructed onto a constitutive expression vector carrying 35S：：GFP, and the recombinant constructs were transferred into Oryza sativajaponica L. cv. Kitaake to investigate the regulatory role of the McPhtgene under phosphorus deficiency. The McPhtgene encodes a protein of 357 amino acids with six transmembrane domains and is located to the mitochondria, and the mRNAtranscripts of the McPht gene are highly accumulated in the shoots of M. crystallinum in response to phosphorus deficiency. However, more mRNA transcripts of the McPht gene were accumulated in the roots of the transgenic rice under phosphorus deficiency. Measurements showed that the transgenic rice demonstrated an enhanced promotion in the root development, the root activities, and phosphate uptake under phosphorus deficiency. Transcriptome sequencing showed that the transgenic rice exhibited total of 198 differentially expressed genes. Of these, total of 154 differentially expressed genes were up-regulated and total 44 genes were down-regulated comparing to the wild type in response to phosphorus deficiency. The selective six genes of the up-regulated differentially expressed genes showed an enhanced increase in mRNA transcripts in response to phosphorus deficiency, however, the transcripts of the mitochondrial carrier protein transporter in rice, a homologous gene of the McPht, in both the transgenic line and the wild type had no obvious differences. Functional enrichment analyses revealed that the most of the up-regulated genes are involved in the cytoplasmic membrane-bounded vesicle, and most of the down-regulated genes are involved in the mitochondrion and cytoplasmic membrane-bounded vesicle. The differentially expressed genes were highly enriched in plant secondary metabolisms and plant-pathogen interaction. These results indicated that the overexpression of the McPht gene might participate in the physiological adaptive modulation of the transgenic rice to phosphorus deficiency by up- or down-regulating the differentially expressed genes.

Functional Analysis of Phosphate Transporter OsPHT4 Family Members in Rice

The transport of phosphate between cytoplasm and subcellular compartments is critical for plant metabolic regulation.We conducted bioinformatic analysis,heterologous expression in yeast,gene expression pattern and subcellular localization analysis to characterize the possible functions of OsPHT4 gene family in rice.Together with the PHT4 genes from higher plants,OsPHT4s can be classified into six distinct groups.OsPHT4;1-OsPHT4;4 are targeted to chloroplasts,and OsPHT4;6-1 and OsPHT4;6-2 are located to Golgi apparatus.OsPHT4 proteins can mediate inorganic phosphate(Pi)transport in yeast.In addition,dynamic transcriptional changes determined by qRT-PCR revealed different expression profiles of OsPHT4 genes in response to phosphate starvation,salicylic acid,abscisic acid and salt stress treatments.These results suggested that OsPHT4 proteins are involved in Pi distribution between the cytoplasm and chloroplast or Golgi apparatus and also involved in stress responses.

Isolation and cloning of the gene encoding high affinity phosphate transporter in rice

High affinity phosphate transporterplays an important role in plantadapting to low phosphorus. Isolationof genes coding this kind of proteinhas attracted worldwide scholars toaccomplish. We aimed to isolate thegene and transfer it to target plants

Complex Regulation of Plant Phosphate Transporters and the Gap between Molecular Mechanisms and Practical Application： What Is Missing？

It has been almost 25 years since the first report of the gene encoding a high-affinity phosphate transporter （PT）, PH084, in yeast. Since then, an increasing number of yeast PH084 homologs as well as other genes encoding proteins with phosphate （Pi） transport activities have been identified and functionally characterized in diverse plant species. Great progress has been made also in deciphering the molecular mechanism underlying the regulation of the abundance and/or activity of these genes and their products. The regulatory genes affect plant Pi homeostasis commonly through direct or indirect regulation of the abundance of PTs at different levels. However, little has been achieved in the use of PTs for developing genetically modified crops with high phosphorus use efficiency （PUE）. This might be a consequence of overemphasizing Pi uptake from the rhizosphere and lack of knowledge about the roles of PTs in Pi transport and recycling within the plant that are required to optimize PUE. Here, we mainly focused on the genes encoding proteins with Pi transport activities and the emerging understanding of their regulation at the transcriptional, posttranscriptional, translational, and post-translational levels. In addition, we propose potential strategies for effective use of PTs in improving plant growth and development.

The Phosphate Transporter PHT4;6 Is a Determinant of Salt Tolerance that Is Localized to the Golgi Apparatus of Arabidopsis

Insertion mutations that disrupt the function of PHT4;6 （At5g44370） cause NaCI hypersensitivity of Arabidopsis seedlings that is characterized by reduced growth of the primary root, enhanced lateral branching, and swelling of root tips. Mutant phenotypes were exacerbated by sucrose, but not by equiosmolar concentrations of mannitol, and attenuated by low inorganic phosphate in the medium. Protein PHT4;6 belongs to the Major Facilitator Superfamily of permeases that shares significant sequence similarity to mammalian type-I Pi transporters and vesicular glutamate transporters, and is a member of the PHT4 family of putative intracellular phosphate transporters of plants. PHT4;6 localizes to the Golgi membrane and transport studies indicate that PHT4;6 facilitates the selective transport of Pi but not of chloride or inorganic anions. Phenotypic similarities with other mutants displaying root swelling suggest that PHT4;6 likely functions in protein N-glycosylation and cell wall biosynthesis, which are essential for salt tolerance. Together, our results indicate that PHT4;6 transports Pi out of the Golgi lumenal space for the re-cycling of the Pi released from glycosylation processes.

Circadian Clock-Regulated Phosphate Transporter PHT4;1 Plays an Important Role in Arabidopsis Defense

The Arabidopsis accelerated cell death 6-1 （acd6-1） mutant shows constitutive defense, cell death, and ex- treme dwarf phenotypes. In a screen for acd6-1 suppressors, we identified a mutant that was disrupted by a T-DNA in the PHOSPHATE TRANSPORTER 4;1 （PHT4;1） gene. The suppressor mutant pht4;1-1 is dominant, expresses truncated PHT4;1 transcripts, and is more susceptible to virulent Pseudomonas syringae strains but not to several avirulent strains. Treat- ment with a salicylic acid （SA） agonist induced a similar level of resistance in Col-0 and pht4;1-1, suggesting that PHT4;1 acts upstream of the SA pathway. Genetic analysis further indicates that PHT4,1 contributes to SID2-dependent and -in- dependent pathways. Transgenic expression of the DNA fragment containing the PHT4;1-1 region or the full-length PHT4;1 gene in wild-type conferred enhanced susceptibility to Pseudomonas infection. Interestingly, expression of PHT4;1 is reg- ulated by the circadian clock. Together, these data suggest that the phosphate transporter PHT4;1 is critical for basal defense and also implicate a potential role of the circadian clock in regulating innate immunity of Arabidopsis.

Characterization of the promoter of phosphate transporter TaPHT1.2 differentially expressed in wheat varieties

TaPHT1.2 is a functional, root predominantly expressed and low phosphate （Pi） inducible high-affinity Pi transporter in wheat, which is more abundant in the roots of P-efficient wheat genotypes （e.g., Xiaoyan 54） than in P-inefficient genotypes （e.g., Jing 411） under both Pi-deficient and Pi-sufficient conditions. To characterize TaPHT1.2 further, we genetically mapped a TaPHT1.2 transporter, TaPHT1.2-D1, on the long arm of chromosome 4D using a recombinant inbred line population derived from Xiaoyan 54 and Jing 411, and isolated a 1,302 bp fragment of the TaPHT1.2-D1 promoter （PrTaPHT1.2-D1） from Xiaoyan 54. TaPHT1.2-D1 shows collinearity with OsPHT1.2 that has previously been reported to mediate the translocation of Pi from roots to shoots. PrTaPHT1.2-D contains a number of Pi-starvation responsive elements, including P1BS, WRKY-binding W-box, and helix-loop-helix-binding elements. PrTaPHT1.2-D1 was then used to drive expression of 13-glucuronidase （GUS） reporter gene in Arabidopsis through Agrobacterium-mediated transformation. Histochemical analysis of transgenic Arabidopsis plants showed that the reporter gene was specifically induced by Pi-starvation and predominantly expressed in the roots. As there is only one SNP between the TaPHT1.2-D1 promoters of Xiaoyan 54 and Jing 411, and this SNP does not exist within the Pi-starvation responsive elements, the differential expression of TaPHT1.2 in Xiaoyan 54 and Jing 411 may not be caused by this SNP.

Cloning,expression and function of phosphate transporter encoded gene in Oryza sativa L.

OsPT6:1,a phosphate transporter encoding gene from the leaf samples of Oryza sativa, was identified through PCR with specifically designed primers.The phylogenetic analysis and the conserved amino acid residue site detection suggested OsPT6:1 a possible high-affinity phosphate transporter encoding gene.In situ hybridization and RT-PCR demonstrated the expression of OsPT6:1 in both roots and leaves.The peak expression signal was observed in mesophyll cells under low phosphorus(P)induction.A homologous recombination study indicated that OsPT6:1 can enhance the Pi uptake efficiency of Pichia pastoris.At the meantime,the introduction of OsPT6:1 was able to complement the Pi uptake function of yeast cells with high-affinity phosphate transporters de- ficient.Those results substantiated our contention that OsPT6:1 encoded a high-affinity phosphate transporter of Oryza sativa.

The Effect of Dietary Nutrient Density on Growth Performance, Physiological Parameters, and Small Intestinal Type IIb Sodium Phosphate Co-transporter Expression in Broilers

A 3 × 4 factorial experiment was con- ducted to determine the effect of dietary nutrient den- sity on growth performance, physiological parame- ters, and small intestinal epithelial phosphate trans- porter expression in broiler chicks fed different dietary nutrient density （DND） and non-phytate phosphorus （NPP） levels. Dietary energy densities used had ME values of 2,850, 2,950, and 3,050 kcal/kg of diet and the NPP levels were 0.35%, 0.40%, 0.45%, and 0.50% within each ME value. Crude protein and essential amino acids levels were maintained propor- tionally to dietary ME levels. Each of the twelve diets was fed to five pens of seven male broiler chicks from 1 to 21 days of age. Broiler growth performance, ser- um physiological parameters, 1-hydroxylase activity in the kidney, type-IIb sodium phosphate co-trans- porter （NaPi-IIb）, and vitamin D receptor （VDR） expression levels in the duodenal mucosa were deter- mined. Our results showed that an increase in dietary nutrient density increased body weight and improved feed conversion. Additionally, serum parathyroid hormone concentration, 1-a-hydroxylase activity in the kidney, NaPi-IIb mRNA expression and VDR protein expression in the duodenal mucosa increased as DND increased. Feed intake and body weight in- creased as NPP levels increased. Serum parathyroid hormone, VDR and NaPi-IIb gene expression also in- creased as the level of NPP in the diet increased at the medium range level. The results of this study suggest that high DND can increase small intestinal type IIb sodium dependent phosphate co-transporter mRNA expression by up-regulating parathyroid hormone and activation of 1-ot-hydroxylase activity which might be in consistent with high growth rate of broilers. Small intestinal NaPi-IIb mRNA expression increases linear- ly only within the range of dietary NPP dosage levels close to the requirement recommended in the broilers feeding standards.