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A purple acid phosphatase plays a role in nodule formation and nitrogen fixation in Astragalus sinicus

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Abstract

The AsPPD1 gene from Astragalus sinicus encodes a purple acid phosphatase. To address the functions of AsPPD1 in legume-rhizobium symbiosis, its expression patterns, enzyme activity, subcellular localization, and phenotypes associated with its over-expression and RNA interference (RNAi) were investigated. The expression of AsPPD1 was up-regulated in roots and nodules after inoculation with rhizobia. Phosphate starvation reduced the levels of AsPPD1 transcripts in roots while increased those levels in nodules. We confirmed the acid phosphatase and phosphodiesterase activities of recombinant AsPPD1 purified from Pichia pastoris, and demonstrated its ability to hydrolyze ADP and ATP in vitro. Subcellular localization showed that AsPPD1 located on the plasma membranes in hairy roots and on the symbiosomes membranes in root nodules. Over-expression of AsPPD1 in hairy roots inhibited nodulation, while its silencing resulted in nodules early senescence and significantly decreased nitrogenase activity. Furthermore, HPLC measurement showed that AsPPD1 overexpression affects the ADP levels in the infected roots and nodules, AsPPD1 silencing affects the ratio of ATP/ADP and the energy charge in nodules, and quantitative observation demonstrated the changes of AsPPD1 transcripts level affected nodule primordia formation. Taken together, it is speculated that AsPPD1 contributes to symbiotic ADP levels and energy charge control, and this is required for effective nodule organogenesis and nitrogen fixation.

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References

  • Annadana S, Schipper B, Beekwilder J, Outchkourov N, Udayakumar M, Jongsma MA (2003) Cloning, functional expression in Pichia pastoris, and purification of potato cystatin and multicystatin. J Biosci Bioeng 95:118–123

    Article  CAS  PubMed  Google Scholar 

  • Antonyuk SV, Olczak M, Olczak T, Ciuraszkiewicz J, Strange RW (2014) The structure of a purple acid phosphatase involved in plant growth and pathogen defence exhibits a novel immunoglobulin-like fold. IUCrJ 1:101–109

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Chen WX, Li GS, Qi YL, Wang ET, Wang HL, Yuan HL, Li L (1991) Rhizobium huakuii sp. nov. isolated from the root nodules of Astragalus sinicus. Int J Syst Bacteriol 41:275–280

    Article  Google Scholar 

  • Chen T, Zhu H, Ke D, Cai K, Wang C, Gou H, Hong Z, Zhang Z (2012) A MAP kinase kinase interacts with SymRK and regulates nodule organogenesis in Lotus japonicus. Plant Cell 24:823–838

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Ching TM, Hedtke S, Russell SA, Evans HJ (1975) Energy state and dinitrogen fixation in soybean nodules of dark-grown plants. Plant Physiol 55:796–798

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Cho HJ, Widholm JM (2002) Improved shoot regeneration protocol for hairy roots of the legume Astragalus sinicus. Plant Cell Tissue Organ Cult 69:259–269

    Article  CAS  Google Scholar 

  • Dionisio G, Brinch-Pedersen H, Welinder KG, Jørgensen M (2011a) Different site-specific N-glycan types in wheat (Triticum aestivum L.) PAP phytase. Phytochemistry 72:1173–1179

    Article  CAS  PubMed  Google Scholar 

  • Dionisio G, Madsen CK, Holm PB, Welinder KG, Jorgensen M, Stoger E, Arcalis E, Brinch-Pedersen H (2011b) Cloning and characterization of purple acid phosphatase phytases from wheat, barley, maize, and rice. Plant Physiol 156:1087–1100

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Furstenau CR, Trentin Dda S, Gossenheimer AN, Ramos DB, Casali EA, Barreto-Chaves ML, Sarkis JJ (2008) Ectonucleotidase activities are altered in serum and platelets of L-NAME-treated rats. Blood Cells Mol Dis 41:223–229

    Article  PubMed  Google Scholar 

  • Govindarajulu M, Kim SY, Libault M, Berg RH, Tanaka K, Stacey G, Taylor CG (2009) GS52 ecto-apyrase plays a critical role during soybean nodulation. Plant Physiol 149:994–1004

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Hardy RWF, Burns RC, Holsten RD (1973) Applications of the acetylene-ethylene assay for measurement of nitrogen fixation. Soil Biol Biochem 5:47–81

    Article  CAS  Google Scholar 

  • Holmquist M, Tessier DC, Cygler M (1997) High-level production of recombinant Geotrichum candidum lipases in yeast Pichia pastoris. Protein Expr Purif 11:35–40

    Article  CAS  PubMed  Google Scholar 

  • Hurley BA, Tran HT, Marty NJ, Park J, Snedden WA, Mullen RT, Plaxton WC (2010) The dual-targeted purple acid phosphatase isozyme AtPAP26 is essential for efficient acclimation of Arabidopsis to nutritional phosphate deprivation. Plant Physiol 153:1112–1122

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Kaida R, Satoh Y, Bulone V, Yamada Y, Kaku T, Hayashi T, Kaneko TS (2009) Activation of beta-glucan synthases by wall-bound purple acid phosphatase in tobacco cells. Plant Physiol 150:1822–1830

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Kaida R, Serada S, Norioka N, Norioka S, Neumetzler L, Pauly M, Sampedro J, Zarra I, Hayashi T, Kaneko TS (2010) Potential role for purple acid phosphatase in the dephosphorylation of wall proteins in tobacco cells. Plant Physiol 153:603–610

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Kuang R, Chan KH, Yeung E, Lim BL (2009) Molecular and biochemical characterization of AtPAP15, a purple acid phosphatase with phytase activity, in Arabidopsis. Plant Physiol 151:199–209

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Lei L, Chen L, Shi X, Li Y, Wang J, Chen D, Xie F, Li Y (2014) A nodule-specific lipid transfer protein AsE246 participates in transport of plant-synthesized lipids to symbiosome membrane and is essential for nodule organogenesis in Chinese milk vetch. Plant Physiol 164:1045–1058

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Li D, Zhu H, Liu K, Liu X, Leggewie G, Udvardi M, Wang D (2002) Purple acid phosphatases of Arabidopsis thaliana. Comparative analysis and differential regulation by phosphate deprivation. J Biol Chem 277:27772–27781

    Article  CAS  PubMed  Google Scholar 

  • Li WY, Shao G, Lam HM (2008a) Ectopic expression of GmPAP3 alleviates oxidative damage caused by salinity and osmotic stresses. New Phytol 178:80–91

    Article  CAS  PubMed  Google Scholar 

  • Li Y, Zhou L, Chen D, Tan X, Lei L, Zhou J (2008b) A nodule-specific plant cysteine proteinase, AsNODF32, is involved in nodule senescence and nitrogen fixation activity of the green manure legume Astragalus sinicus. New Phytol 180:185–192

    Article  CAS  PubMed  Google Scholar 

  • Li C, Gui S, Yang T, Walk T, Wang X, Liao H (2012) Identification of soybean purple acid phosphatase genes and their expression responses to phosphorus availability and symbiosis. Ann Bot 109:275–285

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Liang C, Sun L, Yao Z, Liao H, Tian J (2012) Comparative analysis of PvPAP gene family and their functions in response to phosphorus deficiency in common bean. PLoS One 7:e38106

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Liao H, Wong FL, Phang TH, Cheung MY, Li WY, Shao G, Yan X, Lam HM (2003) GmPAP3, a novel purple acid phosphatase-like gene in soybean induced by NaCl stress but not phosphorus deficiency. Gene 318:103–111

    Article  CAS  PubMed  Google Scholar 

  • Limpens E, Ivanov S, van Esse W, Voets G, Fedorova E, Bisseling T (2009) Medicago N2-fixing symbiosomes acquire the endocytic identity marker Rab7 but delay the acquisition of vacuolar identity. Plant Cell 21:2811–2828

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Liu Y, Ahn JE, Datta S, Salzman RA, Moon J, Huyghues-Despointes B, Pittendrigh B, Murdock LL, Koiwa H, Zhu-Salzman K (2005) Arabidopsis vegetative storage protein is an anti-insect acid phosphatase. Plant Physiol 139:1545–1556

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Nourizad N, Ehn M, Gharizadeh B, Hober S, Nyrén P (2003) Methylotrophic yeast Pichia pastoris as a host for production of ATP-diphosphohydrolase (apyrase) from potato tubers (Solanum tuberosum). Protein Expr Purif 27:229–237

    Article  CAS  PubMed  Google Scholar 

  • Olczak M, Olczak T (2002) Diphosphonucleotide phosphatase/phosphodiesterase from yellow lupin (Lupinus luteus L.) belongs to a novel group of specific metallophosphatases. FEBS Lett 519:159–163

    Article  CAS  PubMed  Google Scholar 

  • Olczak M, Olczak T (2005) Expression and purification of active plant diphosphonucleotide phosphatase/phosphodiesterase from baculovirus-infected insect cells. Protein Expr Purif 39:116–123

    Article  CAS  PubMed  Google Scholar 

  • Olczak M, Kobialka M, Watorek W (2000) Characterization of diphosphonucleotide phosphatase/phosphodiesterase from yellow lupin (Lupinus luteus) seeds. Biochim Biophys Acta 1478:239–247

    Article  CAS  PubMed  Google Scholar 

  • Olczak M, Morawiecka B, Watorek W (2003) Plant purple acid phosphatases—Genes, structures and biological function. Acta Biochim Pol 50:1245–1256

    CAS  PubMed  Google Scholar 

  • Olczak M, Ciuraszkiewicz J, Wojtowicz H, Maszczak D, Olczak T (2009) Diphosphonucleotide phosphatase/phosphodiesterase (PPD1) from yellow lupin (Lupinus luteus L.) contains an iron-manganese center. FEBS Lett 583:3280–3284

    Article  CAS  PubMed  Google Scholar 

  • Oresnik IJ, Layzell DB (1994) Composition and distribution of adenylates in soybean (Glycine max L.) nodule tissue. Plant Physiol 104:217–225

    CAS  PubMed Central  PubMed  Google Scholar 

  • Ott T, van Dongen JT, Gunther C, Krusell L, Desbrosses G, Vigeolas H, Bock V, Czechowski T, Geigenberger P, Udvardi MK (2005) Symbiotic leghemoglobins are crucial for nitrogen fixation in legume root nodules but not for general plant growth and development. Curr Biol 15:531–535

    Article  CAS  PubMed  Google Scholar 

  • Panter S, Thomson R, de Bruxelles G, Laver D, Trevaskis B, Udvardi M (2000) Identification with proteomics of novel proteins associated with the peribacteroid membrane of soybean root nodules. Mol Plant Microbe Interact 13:325–333

    Article  CAS  PubMed  Google Scholar 

  • Peng SE, Wang YB, Wang LH, Chen WN, Lu CY, Fang LS, Chen CS (2010) Proteomic analysis of symbiosome membranes in Cnidaria-dinoflagellate endosymbiosis. Proteomics 10:1002–1016

    Article  CAS  PubMed  Google Scholar 

  • Penheiter AR, Duff SM, Sarath G (1997) Soybean root nodule acid phosphatase. Plant Physiol 114:597–604

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Price GD, Day DA, Gresshoff PM (1987) Rapid isolation of intact peribacteroid envelopes from soybean nodules and demonstration of selective permeability to metabolites. J Plant Physiol 130:157–164

    Article  CAS  Google Scholar 

  • Rips S, Bentley N, Jeong IS, Welch JL, von Schaewen A, Koiwa H (2014) Multiple N-glycans cooperate in the subcellular targeting and functioning of Arabidopsis KORRIGAN1. Plant Cell 26:3792–3808

    Article  CAS  PubMed  Google Scholar 

  • Robinson WD, Carson I, Ying S, Ellis K, Plaxton WC (2012) Eliminating the purple acid phosphatase AtPAP26 in Arabidopsis thaliana delays leaf senescence and impairs phosphorus remobilization. New Phytol 196:1024–1029

    Article  CAS  PubMed  Google Scholar 

  • Sujkowska M, Borucki W, Golinowski M (2006) Localization of acid phosphatase activity in the apoplast of pea (Pisum sativum L.) root nodules grown under phosphorus deficiency. Acta Physiol Plant 28:263–271

    Article  CAS  Google Scholar 

  • Sun F, Carrie C, Law S, Murcha MW, Zhang R, Law YS, Suen PK, Whelan J, Lim BL (2012) AtPAP2 is a tail-anchored protein in the outer membrane of chloroplasts and mitochondria. Plant Signal Behav 7:927–932

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Tian J, Wang C, Zhang Q, He X, Whelan J, Shou H (2012) Overexpression of OsPAP10a, a root-associated acid phosphatase, increased extracellular organic phosphorus utilization in rice. J Integr Plant Biol 54:631–639

    Article  CAS  PubMed  Google Scholar 

  • Upchurch R, Mortenson LE (1980) In vivo energetics and control of nitrogen fixation: changes in the adenylate energy charge and adenosine 5′-diphosphate/adenosine 5′-triphosphate ratio of cells during growth on dinitrogen versus growth on ammonia. J Bacteriol 143:274–284

    CAS  PubMed Central  PubMed  Google Scholar 

  • Wang L, Li Z, Qian W, Guo W, Gao X, Huang L, Wang H, Zhu H, Wu JW, Wang D, Liu D (2011) The Arabidopsis purple acid phosphatase AtPAP10 is predominantly associated with the root surface and plays an important role in plant tolerance to phosphate limitation. Plant Physiol 157:1283–1299

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Xie X, Huang W, Liu F, Tang N, Liu Y, Lin H, Zhao B (2013) Functional analysis of the novel mycorrhiza-specific phosphate transporter AsPT1 and PHT1 family from Astragalus sinicus during the arbuscular mycorrhizal symbiosis. New Phytol 198:836–852

    Article  CAS  PubMed  Google Scholar 

  • Yamamoto M, Tantikanjana T, Nishio T, Nasrallah ME, Nasrallah JB (2014) Site-specific N-glycosylation of the s-locus receptor kinase and its role in the self-incompatibility response of the brassicaceae. Plant Cell 26:4749–4762

    Article  CAS  PubMed  Google Scholar 

  • Zhang W, Gruszewski HA, Chevone BI, Nessler CL (2008) An Arabidopsis purple acid phosphatase with phytase activity increases foliar ascorbate. Plant Physiol 146:431–440

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Zheng SJ, Khrustaleva L, Henken B, Sofiari E, Jacobsen E, Kik C, Krens F (2001) Agrobacterium tumefaciens-mediated transformation of Allium cepa L.: the production of transgenic onions and shallots. Mol Breed 7:101–115

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (No. 31000115, No. 31371549 and No. 31460056) and the Major State Basic Research Development Program of China (973 Program, No. 2010CB126502) and the State Key Laboratory of Agricultural Microbiology (Grant No. AMLKF200909). We are very grateful to Professor Zhongming Zhang for providing pCAMBIA1301-35S-int-T7 for the RNAi experiment.

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Correspondence to Yixing Li or Youguo Li.

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Wang, J., Si, Z., Li, F. et al. A purple acid phosphatase plays a role in nodule formation and nitrogen fixation in Astragalus sinicus . Plant Mol Biol 88, 515–529 (2015). https://doi.org/10.1007/s11103-015-0323-0

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