Intracellular Acid Phosphatase Research Articles

Antibacterial activity and mechanism of combination natural essential oils from oregano and cinnamon against enterotoxigenic Escherichia coli K99

Essential oils from oregano (OEO) and cinnamon (CEO) have been reported an excellent antibacterial activity, but their synergistic antibacterial effect is rarely studied. In this study, the chemical compositions of OEO, CEO and mixed OEO with CEO (MEO) and their antibacterial activities were analyzed, the mode of MEO action against enterotoxigenic Escherichia coli K99 (ETEC K99) were investigated. Results showed that MEO had the synergistic antibacterial effect, which was proven by fractional inhibitory concentration index of 0.375. The inhibition zone diameter measured varying with volume ratio of OEO and CEO showed that the best synergistic effect was at a ratio 2:8. Meanwhile, the antibacterial mechanism of MEO demonstrated that the MEO changed the morphology of ETEC K99 by scanning and transmission electron microscopy, and disrupted the cell membrane integrity that were proved by the leakage of intracellular nucleic acid and lower alkaline phosphatase activity of ETEC K99. In addition, the MEO also inhibited DNA biosynthesis, biofilm formation and scavenged mature biofilm of ETEC K99. These results confirm the synergistic effect on improvement of antibacterial activity and elucidate a variety of mechanisms through which the ETEC K99 was inactivated, and support the application of MEO to against ETEC K99.

The role of manganese in morphogenesis and pathogenesis of the opportunistic fungal pathogen Candida albicans.

Metals such as Fe, Cu, Zn, and Mn are essential trace nutrients for all kingdoms of life, including microbial pathogens and their hosts. During infection, the mammalian host attempts to starve invading microbes of these micronutrients through responses collectively known as nutritional immunity. Nutritional immunity for Zn, Fe and Cu has been well documented for fungal infections; however Mn handling at the host-fungal pathogen interface remains largely unexplored. This work establishes the foundation of fungal resistance against Mn associated nutritional immunity through the characterization of NRAMP divalent metal transporters in the opportunistic fungal pathogen, Candida albicans. Here, we identify C. albicans Smf12 and Smf13 as two NRAMP transporters required for cellular Mn accumulation. Single or combined smf12Δ/Δ and smf13Δ/Δ mutations result in a 10-80 fold reduction in cellular Mn with an additive effect of double mutations and no losses in cellular Cu, Fe or Zn. As a result of low cellular Mn, the mutants exhibit impaired activity of mitochondrial Mn-superoxide dismutase 2 (Sod2) and cytosolic Mn-Sod3 but no defects in cytosolic Cu/Zn-Sod1 activity. Mn is also required for activity of Golgi mannosyltransferases, and smf12Δ/Δ and smf13Δ/Δ mutants show a dramatic loss in cell surface phosphomannan and in glycosylation of proteins, including an intracellular acid phosphatase and a cell wall Cu-only Sod5 that is key for oxidative stress resistance. Importantly, smf12Δ/Δ and smf13Δ/Δ mutants are defective in formation of hyphal filaments, a deficiency rescuable by supplemental Mn. In a disseminated mouse model for candidiasis where kidney is the primary target tissue, we find a marked loss in total kidney Mn during fungal invasion, implying host restriction of Mn. In this model, smf12Δ/Δ and smf13Δ/Δ C. albicans mutants displayed a significant loss in virulence. These studies establish a role for Mn in Candida pathogenesis.

Rice ACID PHOSPHATASE 1 regulates Pi stress adaptation by maintaining intracellular Pi homeostasis.

The concentration and homeostasis of intracellular phosphate (Pi) are crucial for sustaining cell metabolism and growth. During short-term Pi starvation, intracellular Pi is maintained relatively constant at the expense of vacuolar Pi. After the vacuolar stored Pi is exhausted, the plant cells induce the synthesis of intracellular acid phosphatase (APase) to recycle Pi from expendable organic phosphate (Po). In this study, the expression, enzymatic activity and subcellular localization of ACID PHOSPHATASE 1 (OsACP1) were determined. OsACP1 expression is specifically induced in almost all cell types of leaves and roots under Pi stress conditions. OsACP1 encodes an acid phosphatase with broad Po substrates and localizes in the endoplasmic reticulum (ER) and Golgi apparatus (GA). The phylogenic analysis demonstrates that OsACP1 has a similar structure with human acid phosphatase PHOSPHO1. Overexpression or mutation of OsACP1 affected Po degradation and utilization, which further influenced plant growth and productivity under both Pi-sufficient and Pi-deficient conditions. Moreover, overexpression of OsACP1 significantly affected intracellular Pi homeostasis and Pi starvation signalling. We concluded that OsACP1 is an active acid phosphatase that regulates rice growth under Pi stress conditions by recycling Pi from Po in the ER and GA.

Potato root and leaf phosphatase activity in response to P deprivation

AbstractBackgroundPlants are not able to take up organically bound phosphorus (P) before it is hydrolyzed by extracellular phosphatases while intracellular phosphatases play a role in P remobilization.AimsThe aim of this study was to evaluate intra‐ and extracellular acid phosphatase activity as well as root and shoot growth of two modern starch potato cultivars at different levels of P deprivation.MethodsIn vitro propagated potato plantlets (cultivars Kuba and Cardoso) were grown in sand and fertigated with five different P concentrations, ranging from 1 to 0.05 mM P. Growth parameters, plant P concentrations, and acid phosphatase activities were determined after 40 days.ResultsShoot and root biomass decreased gradually with decreasing P supply, while the root‐to‐shoot ratio increased. Shoot P concentration decreased steadily, but root P concentration remained constant at P levels below 0.5 mM. Root‐associated and root intracellular acid phosphatase activity increased in both cultivars with rising levels of P deprivation. The activity of acid phosphatases in old but not in young leaves was increased with P deprivation. Intracellular acid phosphatase activity was genotype dependent, with higher activity in the cultivar Kuba than in Cardoso.ConclusionsOur results point to a tissue specific regulation of acid phosphatase activity depending on plant P status and potato genotype. Increasing root‐associated acid phosphatase activities may help P starved potato plants to ensure a sufficient P nutrition. Further research is needed to unravel the roles of leaf intracellular acid phosphatases in potato.

A feasible self-assembled near-infrared fluorescence sensor for acid phosphatase detection and cell imaging.

The single signal amplification strategy is significant for detecting various disease biomarkers but is restricted by its limited accuracy. The multi-signal and multi-mode methods have overcome this deficiency. Acid phosphatase (ACP) is an important intracellular enzyme but one-step cell imaging material-based probes are scarce for ACP. Herein, we designed a one-step self-assembled polymer probe using neutral red (NR), modified-(pyridoxal-5'-phosphate (PLP)) and Eu3+. The polymer exhibited non-emission and excellent stability. Upon the catalytic hydrolysis reaction of ACP, the polymer exhibited two strong fluorescence signals at 373 nm and 613 nm and an appreciable decline of absorbance at 395 nm. The probe has excellent selectivity and higher sensitivity with a limit of detection as low as 0.02 mU mL-1. It possesses favorable biocompatibility and has been successfully used to detect and image intracellular ACP in several living cells.

Measurement of Acid Ecto-phosphatase Activity in Live Leishmania donovani Parasites

Acid ecto-phosphatases are enzymes that hydrolyze phosphomonoesters in the acidic pH range with their active sites facing the extacellular medium. Their activities can be measured in living cells. In bacteria and protozoan pathogens, acid ecto-phosphatases have been associated with the survival of intracellular pathogens within phagocytes through inhibition of the respiratory burst, suggesting that they act as virulence factors. Extracellular acid phosphatase activity in Leishmania (L.) donovani has been associated with the degree of promastigote virulence/infectivity. The levels of acid ecto-phosphatase activity in different Leishmania sp or even strains of the same species vary and this has been linked to their virulence. It may also be related to their ability to survive and multiply in the insect host. Acid phosphatase enzymatic activity can be measured in crude membrane fractions and in membrane fractions enriched in plasma membrane, however, in these cases, the intracellular acid phosphatases, mainly localized in lysosomes, contribute to the final result. Therefore, measuring phosphatase activity at the surface of live cells in acidic pH range is the only accurate way to measure acid ecto-phosphatase activity. This assay is performed at 25 °C or 37 °C for 30 min using as substrate the generic phosphatase substrate p-nitrophenyl phosphate (pNPP), in a citrate buffer, with or without sodium tartrate (L(+)-tartaric acid), as histidine acid phosphatases are classified according to their sensitivity to tartate inhibition. The steps of the protocol consist of pelleting cells in suspension, in this case Leishmania promastigotes, washing twice with HEPES buffer, resuspending the cells in the substrate reaction mixture and terminating the reaction by the addition of 0.5 N NaOH. The cells are removed by centrifugation and the absorbance of the reaction product (p-nitrophenolate=pNP) in the supernatant is measured at 405 nm. The enzymatic activity (A405 values) is normalized for the mean number of cells/ml used for each independent experiment.

Mechanisms of oat (Avena sativa L.) acclimation to phosphate deficiency.

BackgroundDeficiency of available forms of phosphorus is common in most soils and causes reduction of crop plants growth and yield. Recently, model plants responses to phosphate (Pi) deficiency have been intensively studied. However, acclimation mechanisms of cereals like oat (Avena sativa L.), to low Pi stress remains not fully understood. Oat plants have been usually cultured on poor soils, with a low nutrient content, but their responses to such conditions are not well known, therefore the main goal of the study was to investigate the mechanisms that enable oat plants to grow under low Pi conditions.MethodsFour oat cultivars (A. sativa, cv. Arab, Krezus, Rajtar and Szakal) were grown for three weeks in a nutrient media with various P sources: inorganic—KH2PO4 (control), organic—phytate (PA) and with no phosphate (−P). The effects of Pi deficiency on the level of P, oat growth parameters, intensity of photosynthesis, plant productivity, root exudation ability, localization, activity and isoforms of acid phosphatases, enzymes involved in Pi mobilization, were estimated. In addition, the effect of mycorrhization on plant growth was also observed.ResultsAll studied oat cultivars grown on Pi-deficient media had significantly decreased Pi content in the tissues. Pi deficiency caused inhibition of shoot growth, but generally it did not affect root elongation; root diameter was decreased, root/shoot ratios increased, whereas PA plants showed a similar growth to control. Photosynthesis rate and productivity parameters decreased under low Pi nutrition, however, sugar content generally increased. Studied oat cultivars did not respond to low Pi via increased exudation of carboxylates from the roots, as pH changes in the growth media were not observed. Pi starvation significantly increased the activity of extracellular and intracellular acid phosphatases (APases) in comparison to the control plants. Three major APase isoforms were detected in oat tissues and the isoform pattern was similar in all studied conditions, usually with a higher level of one of the isoforms under Pi starvation. Generally no significant effects of mycorrhizal colonization on growth of oat cultivars were observed.DiscussionWe postulated that acid phosphatases played the most important role in oat cultivars acclimation to Pi deficiency, especially extracellular enzymes involved in Pi acquisition from soil organic P esters. These APases are mainly located in the epidermis of young roots, and may be released to the rhizosphere. On the other hand, intracellular APases could be involved in fast Pi remobilization from internal sources. Our study showed that oat, in contrast to other plants, can use phytates as the sole source of P. The studied oat cultivars demonstrated similar acclimation mechanisms to Pi deficiency, however, depending on stress level, they can use different pools of acid phosphatases.

Acid Phosphatases Activity and Growth of Barley, Oat, Rye and Wheat Plants as Affected by Pi Deficiency

Objective: The influence of phosphorus deficit on the growth of plants and acid phosphatases activity in leaves and roots of barley seedlings (Hordeum vulgare L.), as well as oat (Avena sativa L.), rye (Secale cereale L.) and wheat plants (Triticum vulgare L.) was studied. Method: Plants were cultured three weeks in a nutrient media: complete (control, +P) or without phosphorus (-P). The growth on -P medium significantly affected the inorganic phosphate (Pi) content in plants tissues. Pi deficit decreased shoots growth but ratio of root/shoot was higher for -P plants when compared to control. The root elongation was enhanced under Pi deficiency - in -P oat and barley more intensive elongation was observed than in other plants. On the other hand, inhibition of shoot growth was more pronounced for -P rye and wheat. Pi-deficient plants showed higher activity of acid phosphatases in tissue extracts and in exudates from roots than +P plants. Result: Extracellular acid phosphatases activity increased the most for -P rye and wheat plants. Acid phosphatases secretion was intensive in growing parts of Pi-deficient roots. The activity of enzymes secreted by -P roots of all studied plants was higher than intracellular acid phosphatases. Conclusion: Our results indicated that wheat is more sensitive to the Pi deficiency at the early stage of growth than other plants, whereas oat is rather resistant to Pi deficit. The results suggested that acid phosphatases played an important role in acclimation of studied crop plants to moderate Pi deficiency.

OsPAP26 Encodes a Major Purple Acid Phosphatase and Regulates Phosphate Remobilization in Rice.

During phosphate (Pi) starvation or leaf senescence, the accumulation of intracellular and extracellular purple acid phosphatases (PAPs) increases in plants in order to scavenge organic phosphorus (P). In this study, we demonstrated that a PAP-encoding gene in rice, OsPAP26, is constitutively expressed in all tissues. While the abundance of OsPAP26 transcript is not affected by Pi supply, it is up-regulated during leaf senescence. Furthermore, Pi deprivation and leaf senescence greatly increased the abundance of OsPAP26 protein. Overexpression or RNA interference (RNAi) of OsPAP26 in transgenic rice significantly increased or reduced APase activities, respectively, in leaves, roots and growth medium. Compared with wild-type (WT) plants, Pi concentrations of OsPAP26-overexpressing plants increased in the non-senescing leaves and decreased in the senescing leaves. The increased remobilization of Pi from the senescing leaves to non-senescing leaves in the OsPAP26-overexpressing plants resulted in better growth performance when plants were grown in Pi-depleted condition. In contrast, OsPAP26-RNAi plants retained more Pi in the senescing leaves, and were more sensitive to Pi starvation stress. OsPAP26 was found to localize to the apoplast of rice cells. Western blot analysis of protein extracts from callus growth medium confirmed that OsPAP26 is a secreted PAP. OsPAP26-overexpressing plants were capable of converting more ATP into inorganic Pi in the growth medium, which further supported the potential role of OsPAP26 in utilizing organic P in the rhizosphere. In summary, we concluded that OsPAP26 performs dual functions in plants: Pi remobilization from senescing to non-senescing leaves; and organic P utilization.

PUB059 Role of Alveolar Macrophages' Molecular Antennas in the Pathogenesis of Lung Cancer

The internal sources of alpha-emitting radionuclides originated from tobacco smoke or inhaled contaminated aerosol, induce the low-slow dose effects in pulmonary tissue. The health effects may not appear for years. The metabolic, ionic and enzyme properties of alveolar macrophages (AM), represent in situ biochemical, immunological and signaling processes. Apoptotic regulation is important for tissue remodeling. The study was performed in order to elucidate some of the mechanisms included in the pathogenesis of lung cancer. Bronchoalveolar lavage (BAL) was performed in a group of smokers whose average smoking exposure (pack-years) values were < 2, and patients with non-small cell lung cancer (NSCLC) > 40, as well as in the control group of nonsmokers. Differential cell counting and cytochemical analysis of the BAL specimens for intracellular iron content, lipids, glycogen, nonspecific esterases, and acid phosphatase (ACP) were performed by light microscopy. Apoptosis detection was performed by using the TUNEL in situ cytochemical method. Iron content in AM is gradually increasing in smokers, as well as in patients with NSCLC, (p<0.05) in comparison with non-smokers. There were no significant differences for ACP, nonspecific esterases, glycogen and lipid content in AM between the investigated groups. Apoptosis is gradually increasing in smokers and NSCLC group in comparison with nonsmokers. Apoptotic clearance of free apoptotic bodies by AM phagocytosis is increased in smokers in comparison with nonsmokers, but in the NSCLC group, AC is significantly decreased both, in comparison with nonsmokers, as well as control smokers (p<0.05). AM possess iron centers in their enzymes. The regulation of gene expression for ACP depends on iron. The binuclear iron center of ACP is included in the generation of reactive oxygen species. During the prolonged tissue injury, AM gather iron which can itself contribute to the process of cell damage, as an ionic part of molecular antennas (biomolecules, including enzymes with metal ion surrounded by biopolymer chains). Ferroptosis could be an iron-dependent form of non-apoptotic death of AM, without cell energy depletion. Alpha-emitting internal sources of radiation may trigger random and irregular tissue damage and repair. Oxygen diffusion limit increases for the population of proliferating cells. Their metabolic phenotype changes from glycolytic to lipogenic, with the further transformation of cells to the initiation of neoplastic lesion in the prolonged course. The changed biological properties of AM could be of importance as a utility for biological dosimetry purposes.

The purple acid phosphatase GmPAP21 enhances internal phosphorus utilization and possibly plays a role in symbiosis with rhizobia in soybean.

Induction of secreted and intracellular purple acid phosphatases (PAPs; EC 3.1.3.2) is widely recognized as an adaptation of plants to phosphorus (P) deficiency. The secretion of PAPs plays important roles in P acquisition. However, little is known about the functions of intracellular PAP in plants and nodules. In this study, we identified a novel PAP gene GmPAP21 in soybean. Expression of GmPAP21 was induced by P limitation in nodules, roots and old leaves, and increased in roots with increasing duration of P starvation. Furthermore, the induction of GmPAP21 in nodules and roots was more intensive than in leaves in both P-efficient genotype HN89 and P-inefficient genotype HN112 in response to P starvation, and the relative expression in the leaves and nodules of HN89 was significantly greater than that of HN112 after P deficiency treatment. Further functional analyses showed that over-expressing GmPAP21 significantly enhanced both acid phosphatase activity and growth performance of hairy roots under P starvation condition, indicating that GmPAP21 plays an important role in P utilization. Moreover, GUS expression driven by GmPAP21 promoter was shown in the nodules besides roots. Overexpression of GmPAP21 in transgenic soybean significantly inhibited nodule growth, and thereby affected plant growth after inoculation with rhizobia. This suggests that GmPAP21 is also possibly involved in regulating P metabolism in nodules. Taken together, our results suggest that GmPAP21 is a novel plant PAP that functions in the adaptation of soybean to P starvation, possibly through its involvement in P recycling in plants and P metabolism in nodules.

Insights into the physiological responses of the facultative halophyte Aeluropus littoralis to the combined effects of salinity and phosphorus availability

In this work, we investigate the physiological responses to P deficiency (5μM KH2PO4=D), salt stress (400mM NaCl=C+S), and their combination (D+S) on the facultative halophyte Aeluropus littoralis to understand how plants adapt to these combined stresses. When individually applied, both P deficiency and salinity significantly restricted whole plant growth, with a more marked effect of the latter stress. However, the effects of the two stresses were not additive in plant biomass production since the response of plants to combined salinity and P deficiency was similar to that of plants grown under salt stress alone. In addition the observed features under salinity alone are kept when plants are simultaneously subjected to the combined effects of salinity and P deficiency such as biomass partitioning; the synthesis of proline and the K+/Na+ selectivity ratio. Thus, increasing P availability under saline conditions has no significant effect on salt tolerance in this species.Plants cultivated under the combined effects of salinity and P deficiency exhibited the lowest leaf water potential. This trend was associated with a high accumulation of Na+, Cl− and proline in shoots of salt treated plants suggesting the involvement of these solutes in osmotic adjustment. Proline could be involved in other physiological processes such as free radical scavenging. Furthermore, salinity has no significant effect on phosphorus acquisition when combined with a low P supply and it significantly decreased this parameter when combined with a sufficient P supply. This fact was probably due to salt’s effect on P transporters. In addition, shoot soluble sugars accumulation under both P deficiency treatments with and without salt likely play an important role in the adaptation of A. littoralis plants to P shortage applied alone or combined with salinity.Moreover, there was a strong correlation between shoot and root intracellular acid phosphatase activity and phosphorus use efficiency which strengthens the assumption that intracellular acid phosphatase enzymes are involved in P remobilization in this species. Finally, our results showed that P availability has no significant effect on salt excretion in A. littorlais which suggests that independently of the P status in the plant, excretion remains priority over other functions requiring energy such as growth. This result could also indicate that salt excretion is not energy-dependent in this species.

The maize (Zea mays ssp. mays var. B73) genome encodes 33 members of the purple acid phosphatase family.

Purple acid phosphatases (PAPs) play an important role in plant phosphorus nutrition, both by liberating phosphorus from organic sources in the soil and by modulating distribution within the plant throughout growth and development. Furthermore, members of the PAP protein family have been implicated in a broader role in plant mineral homeostasis, stress responses and development. We have identified 33 candidate PAP encoding gene models in the maize (Zea mays ssp. mays var. B73) reference genome. The maize Pap family includes a clear single-copy ortholog of the Arabidopsis gene AtPAP26, shown previously to encode both major intracellular and secreted acid phosphatase activities. Certain groups of PAPs present in Arabidopsis, however, are absent in maize, while the maize family contains a number of expansions, including a distinct radiation not present in Arabidopsis. Analysis of RNA-sequencing based transcriptome data revealed accumulation of maize Pap transcripts in multiple plant tissues at multiple stages of development, and increased accumulation of specific transcripts under low phosphorus availability. These data suggest the maize PAP family as a whole to have broad significance throughout the plant life cycle, while highlighting potential functional specialization of individual family members.

Effect of short-term phosphate starvation on acid phosphatase activity of Carpinus pubescens and Eurycorymbus cavalerei

The objective of this research was to test the effect of phosphate starvation on the plant growth and acid phosphatase activity of Carpinus pubescens and Eurycorymbus cavalerei. Growth parameters of both trees with −P and +P exhibited little change. The Pi content in leaves of both Pi-free trees decreased to about 65–70% of the control after cultivation for two to three weeks. Root Pi content of both Pi deprivation trees were by 30–45% lower than that of the Pi supply. Phosphate starvation significantly increased both intracellular and extracellular acid phosphatase activities. Intracellular and extracellular acid phosphatase activities were negatively correlated with Pi content in roots, while intracellular acid phosphatase activities were significantly positively correlated with Pi content in leaves under Pi starvation. These results indicated that excreted acid phosphatase mainly released Pi from organic phosphorus compounds, while intracellular acid phosphatase in leaves regulated the redistribution of Pi in tissues for adaptation to a calcareous soil with a low phosphate level.

Senescence-inducible cell wall and intracellular purple acid phosphatases: implications for phosphorus remobilization in Hakea prostrata (Proteaceae) and Arabidopsis thaliana (Brassicaceae).

Despite its agronomic importance, the metabolic networks mediating phosphorus (P) remobilization during plant senescence are poorly understood. Highly efficient P remobilization (~85%) from senescing leaves and proteoid roots of harsh hakea (Hakea prostrata), a native 'extremophile' plant of south-western Australia, was linked with striking up-regulation of cell wall-localized and intracellular acid phosphatase (APase) and RNase activities. Non-denaturing PAGE followed by in-gel APase activity staining revealed senescence-inducible 120kDa and 60kDa intracellular APase isoforms, whereas only the 120kDa isoform was detected in corresponding cell wall fractions. Kinetic and immunological properties of the 120kDa and 60kDa APases partially purified from senescing leaves indicated that they are purple acid phosphatases (PAPs). Results obtained with cell wall-targeted hydrolases of harsh hakea were corroborated using Arabidopsis thaliana in which an ~200% increase in cell wall APase activity during leaf senescence was paralleled by accumulation of immunoreactive 55kDa AtPAP26 polypeptides. Senescing leaves of an atpap26 T-DNA insertion mutant displayed a >90% decrease in cell wall APase activity. Previous research established that senescing leaves of atpap26 plants exhibited a similar reduction in intracellular (vacuolar) APase activity, while displaying markedly impaired P remobilization efficiency and delayed senescence. It is hypothesized that up-regulation and dual targeting of PAPs and RNases to the cell wall and vacuolar compartments make a crucial contribution to highly efficient P remobilization that dominates the P metabolism of senescing tissues of harsh hakea and Arabidopsis. To the best of the authors' knowledge, the apparent contribution of cell wall-targeted hydrolases to remobilizing key macronutrients such as P during senescence has not been previously suggested.

Actividad de las fosfatasas ácidas y alcalinas (extracelulares e intracelulares) en hongos de la rizosfera de &lt;i&gt;Arachis hypogaea&lt;/i&gt; (Papiloneaceae

The potential role of the fungi, isolated from the peanut rhizosphere, in the production of extracellular and intracellular acid and alkaline phosphatase, was evaluated in vitro. Acid and alkaline extracellular phosphatases showed the highest activities, and the Penicillium species were the most efficient in their production. The correlation analysis showed that extracellular acid and intracellular acid phosphatase produced by Aspergillus niger A. terreus, Penicillium sp. y P. brevicompactum were negatively correlated; while the extracellular and intracellular phosphatase activities, were positively correlated. The extracellular acid phosphatase activities produced in vitro by majority of fungi assayed, were not correlated with the acid phosphatase activity present in the peanut soil rhizosphere. Nevertheless, the extracellular alkaline phosphatase activities produced in vitro, were negatively correlated with the extracellular alkaline phosphatase activities present in the rhizosphere. The ability of phosphatase production by fungi isolated from peanut rhizosphere suggests they have great potential to contribute to the P mineralization in this zone.

Spatial distribution and expression of intracellular and extracellular acid phosphatases of cluster roots at different developmental stages in white lupin

Acid phosphatases (APases) play a key role in phosphorus (P) acquisition and recycling in plants. White lupin (Lupinus albus L.) forms cluster roots (CRs) and produces large amounts of APases under P deficiency. However, the relationships between the activity of intracellular and extracellular APases (EC 3.1.3.2) and CR development are not fully understood. Here, comparative studies were conducted to examine the spatial variation pattern of APase activity during CR development using the enzyme-labelled fluorescence-97 (ELF-97) and the p-nitrophenyl phosphate methods. The activity of intracellular and extracellular APases was significantly enhanced under P deficiency in the non-CRs and CRs at different developmental stages. These two APases exhibited different spatial distribution patterns during CR development, and these distribution patterns were highly modified by P deficiency. The activity of extracellular APase increased steadily with CR development from meristematic, juvenile, mature to senescent stages under P deficiency. In comparison, P deficiency-induced increase in the activity of intracellular APase remained relatively constant during CR development. Increased activity of intracellular and extracellular APases was associated with enhanced expression of LaSAP1 encoding intracellular APase and LaSAP2 encoding extracellular APase. The expression levels of these two genes were significantly higher at transcriptional level in both mature and senescent CRs. Taken together, these findings demonstrate that both activity and gene expression of intracellular or extracellular APases exhibit a differential response pattern during CR development, depending on root types, CR developmental stages and P supply. Simultaneous in situ determination of intracellular and extracellular APase activity has proved to be an effective approach for studying spatial variation of APases during CR development.

Overexpression of OsPAP10a, A Root‐Associated Acid Phosphatase, Increased Extracellular Organic Phosphorus Utilization in Rice

Phosphorus (P) deficiency is a major limitation for plant growth and development. Among the wide set of responses to cope with low soil P, plants increase their level of intracellular and secreted acid phosphatases (APases), which helps to catalyze inorganic phosphate (Pi) hydrolysis from organo-phosphates. In this study we characterized the rice (Oryza sativa) purple acid phosphatase 10a (OsPAP10a). OsPAP10a belongs to group Ia of purple acid phosphatases (PAPs), and clusters with the principal secreted PAPs in a variety of plant species including Arabidopsis. The transcript abundance of OsPAP10a is specifically induced by Pi deficiency and is controlled by OsPHR2, the central transcription factor controlling Pi homeostasis. In gel activity assays of root and shoot protein extracts, it was revealed that OsPAP10a is a major acid phosphatase isoform induced by Pi starvation. Constitutive overexpression of OsPAP10a results in a significant increase of phosphatase activity in both shoot and root protein extracts. In vivo root 5-bromo-4-chloro-3-indolyl-phosphate (BCIP) assays and activity measurements on external media showed that OsPAP10a is a root-associated APase. Furthermore, overexpression of OsPAP10a significantly improved ATP hydrolysis and utilization compared with wild type plants. These results indicate that OsPAP10a can potentially be used for crop breeding to improve the efficiency of P use.

Salinity a significant environmental factor forVibrio harveyivirulence inFenneropenaeus indicus

The effect of ambient salinity on the haemolymph variables of Fenneropenaeus indicus and its susceptibility to Vibrio harveyi infection under salinity stress has been studied. Adult shrimps were acclimated to 5‰ (hypo osmotic), 25‰ (iso osmotic) and 35‰ (hyper osmotic) salinity levels and the animals were injected with a mid logarithmic culture of V. harveyi at sub lethal level and haemolymph parameters were analysed. Haemolymph proteins, intracellular superoxide anion production, phenoloxidase (PO), alkaline phosphatase (ALP) and acid phosphatase (ACP) activity were found to be at elevated level both at 5‰ and 35‰ post challenge. The haematological responses showed a progressive increase (P < 0.05) up to post challenge day 5 (PCD 5) followed by a considerable decline at all salinities with the lowest being at 35‰. The alterations in the variables were higher in shrimps held at 5‰. However, the V. harveyi infection was severe in animals held at 35‰. The reduction in the parameters could be correlated with the decrease in survival rate of shrimps at 35‰ with a concurrent increase in V. harveyi at this salinity. Multiple regression analysis revealed that ACP (P < 0.001), haemocyte protein HCP (P < 0.001) and PO (P < 0.05) could explain 91% variability in the shrimp survival. These parameters may be used as effective shrimp health indicators. It is evident from the study that ambient salinity alters the haemolymph variables, modulates the virulence in V. harveyi and makes the shrimps more vulnerable to infection at higher salinity. The virulence of V. harveyi is increased at 35‰ salinity as being evidenced from the high mortality at this salinity. The study emphasizes the importance of salinity as an important environmental factor both in terms of host susceptibility and virulence of the pathogen.

High-temperature optima phosphatases from the cold-tolerant Arctic fungusPenicillium citrinum

Fifty-six fungal isolates from Arctic soils were subjected to primary screening for their ability to solubilize insoluble inorganic phosphate. Nine of the isolates were further analysed quantitatively for phosphatase production using para- nitrophenylphosphate as substrate. Amongst these, a cold-tolerant fungus, Penicillium citrinum strain PG162 was found to be the best producer of intracellular acid phosphatase. Further characterization of the enzyme showed that it is most active in the temperature range of 40-60°C and pH range of 4.2-4.8. The dried enzyme extract is stable at a temperature of up to 50°C for at least 1 h. Its activity is affected by presence of metal ions. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS–PAGE) analysis suggests the molecular weight of the enzyme to be between 20 and 29 kDa. The present study is important with respect to our understanding of the kind of enzymatic reactions that take place in the polar microbes, and the extent to which their activity is sustained. Keywords: acid phosphatase; enzyme; para-nitrophenylphosphate; Pikovskaya medium; psychrotolerant; SDSPAGE (Published: 9 August 2012) Citation: Polar Research 2012, 31 , 11105, http://dx.doi.org/10.3402/polar.v31i0.11105