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Pea lectin is correctly processed, stable and active in leaves of transgenic potato plants

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Abstract

A gene encoding the preproprotein of the pea (Pisum sativum) lectin was expressed in transgenic potato plants using a cauliflower mosaic virus (CaMV) 35S promoter or a tobacco ribulose bisphosphate carboxylase small subunit (ssRubisco) promoter. Presence of the pea lectin to levels greater than 1% of total soluble leaf protein was detected by radioimmunoassay (RIA). The pattern of expression derived from the two promoters was established using both RIA and a squash-blot immunolocalisation technique. Western blotting demonstrated that the preproprotein was correctly processed, generating α and β subunits that assembled to give an isolectin form observed in pea seeds and roots. It was also found that the haemagglutination activity and specificity of pea lectin synthesised in transgenic potato leaves was comparable to purified lectin from pea cotyledons.

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References

  1. Allen AK, Neuberger A: Potato lectins. In: Ginsberg V (ed), Methods in Enzymology, vol 50, pp. 340–345. Academic Press, New York (1978).

    Google Scholar 

  2. Beachy RN, Cheu ZL, Horsch RB, Rogers SG, Hoffmann NJ, Fraley RT: Accumulation and assembly of soybean β-conglycinin in seeds of transformed petunia plants. EMBO J 4: 3047–3053 (1985).

    Google Scholar 

  3. Begbie R, King TP: The interaction of dietary lectin with porcine small intestine and the production of lectin specific antibodies. In: Bog-Hansen TC, Boboritch J (eds) Lectins: Biology, Biochemistry, Clinical Biochemistry, vol 5, pp. 15–27. Walter de Gruyter, Berlin (1985).

    Google Scholar 

  4. Bevan M: BinaryAgrobacterium vectors for plant transformation. Nucl Acids Res 12: 8711–21 (1984).

    Google Scholar 

  5. Boulter D. In: Magnien E (ed) Final report in Biomolecular Engineering in the European Community: Achievements in the Research Programme (1982–1986), pp. 715–725. Martinus Nijhoff, The Hague (1986).

    Google Scholar 

  6. Boulter D, Edwards GA, Gatehouse AMR, Gatehouse JA, Hilder VA: Additive protective effects of different plant-derived insect resistance genes in transgenic tobacco plants. Crop Prot 9: 351–354 (1990).

    Google Scholar 

  7. Bradford M: A rapid and sensitive method for the quantitation of microgram quantities of protein using the principle of dye-binding. Analyt Biochem 72: 248–254 (1976).

    Google Scholar 

  8. Czernilofsky AP, Hain R, Herrera-Estrella L, Lorz H, Goyvaerts E, Baker BJ, Schell J: Fate of selectable marker DNA integrated into the genome ofNicotiana tabacum. DNA 5: 101–113 (1986).

    Google Scholar 

  9. Dellaporta SL, Wood J, Hicks JB: A plant DNA mini preparation: version II. Plant Mol Biol Rep 1: 19–21 (1993).

    Google Scholar 

  10. Diaz CL, Hosselet M, Logman GJJ, van Driessche E, Lugtenberg BJ, Kijne JW: Distribution of glucose/mannose-specific isolectins in pea (Pisum sativum L.) seedlings. Planta 181: 451–461 (1990).

    Google Scholar 

  11. Diaz CL, Melchers LS, Hooykaas PJJ, Lugtenberg BJJ, Kijne JW: Root lectin as a determinant of host-plant specificity in theRhizobium-legume symbiosis. Nature 338: 579–581 (1989).

    Google Scholar 

  12. Diaz CL, van Spronsen PC, Bakhuisen R, Logman GL, Lugtenberg EJ, Kijne JW: Correlation between infection ofRhizobium leguminosarum and lectin on the surface ofPisum sativum L. roots. Planta 168: 350–359 (1986).

    Google Scholar 

  13. Edwards GA: Plant transformation using anAgrobacterium tumefaciens Ti-plasmid vector system. Ph.D. Thesis, University of Durham, UK (1988).

  14. Ellis JR, Shirsat AH, Hepher A, Yarwood JN, Gatehouse JA, Croy RRD, Boulter D: Tissue-specific expression of a pea legumin gene in seeds ofNicotiana plumbaginifolia. Plant Mol Biol 10: 203–214 (1988).

    Google Scholar 

  15. Etzler ME. Plant lectins: molecular and biological aspects. Ann Rev Plant Physiol 36: 209–234 (1985).

    Google Scholar 

  16. Gatehouse JA, Brown D, Evans IM, Gatehouse LN, Jobes D, Preston P, Croy RRD: Sequence of the seed lectin gene from pea (Pisum sativum L.). Nucl Acids Res 15: 7642 (1987).

    Google Scholar 

  17. Gatehouse JA, Boulter D: Isolation and properties of a lectin from the roots ofPisum sativum (garden pea). Physiol Plant 49: 437–442 (1980).

    Google Scholar 

  18. Gatehouse JA, Evans IM, Croy RRD, Boulter D: Differential expression of seed genes during legume seed development. Phil Trans R Soc Lond B 314: 341–500 (1986).

    Google Scholar 

  19. Goldsbrough PB, Gelvin SB, Larkins BA: Expression of maize zien genes in transformed sunflower cells. Mol Gen Genet 202: 374–381 (1986).

    Google Scholar 

  20. Grant G: Antinutritional effects of dietary lectins. Aspects Appl Biol 19: 51–74 (1989).

    Google Scholar 

  21. Hall TC, Buchbinder BV, Pyne JW, Sun SM, Bliss FM: Messenger RNA for G1 protein of french bean seeds: cell-free translation and product characterisation. Proc Natl Acad Sci USA 75: 3196–3200 (1978).

    Google Scholar 

  22. Higgins TJV, Chandler PM, Zurawaski G, Button SC, Spencer D: The synthesis and primary structure of pea seed lectin. J Biol Chem 258: 9544–9549 (1983).

    Google Scholar 

  23. Higgins TJV, Chrispeels MJ, Chandler PM, Spencer D: Intracellular sites of synthesis and processing of lectin in pea cotyledons. J Biol Chem 258: 9550–9552 (1983).

    Google Scholar 

  24. Jacobsen E, Templelaar MJ, Bijmout EW: Ploidy levels in leaf callus and regenerated plants ofSolanum tuberosum determined by cytophotometric measurements of protoplasts. Theor Appl Genet 65: 113–118 (1983).

    Google Scholar 

  25. Jahn R, Schiebler W, Greengard P: A quantitative dotimmunobinding assay for proteins using nitrocellulose. Proc Natl Acad Sci USA 81: 1684–1687 (1984).

    Google Scholar 

  26. Johnson DA, Gautsch JW, Sportsman JR, Elder JH: Improved technique utilizing nonfat dry milk for analysis of proteins and nucleic acids transfered to nitrocellulose. Gene Anal Tech 1: 3–8 (1984).

    Google Scholar 

  27. Kijne JW, Van der Schaal IAM, Diaz CL, Van Iren F: Mannose specific lectins and the recognition of pea roots byRhizobium leguminosarum. In: Bog-Hansen TC, Spengler GA (eds) Lectins: Biology, Biochemistry, Clinical Biochemistry, vol 3, pp. 521–529. Walter de Gruyter, Berlin (1983).

    Google Scholar 

  28. Kyhse-Anderson J: Electroblotting of multiple gels: a simple apparatus without buffer tank for rapid transfer of proteins from polyacrylamide to nitrocellulose. J Biochem Biophys Meth 10: 203–209 (1984).

    Google Scholar 

  29. Lawton MA, Tierney MA, Nakamura I, Anderson E, Komeda Y, Dube P, Hoffman N, Fraley RT, Beachy RN: Expression of a soybean β-conglycinin gene under the control of the cauliflower mosaic virus 35S and 19S promoters in transformed petunia tissues. Plant Mol Biol 9: 315–324 (1987).

    Google Scholar 

  30. Lis H, Sharon N: Lectins as molecules and tools. Ann Rev Biochem 55: 35–68 (1986).

    Google Scholar 

  31. Maniatis T, Fritsch EF, Sambrook J: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982).

    Google Scholar 

  32. McMarsters GK, Carmichael GG: Analysis of single- and double-stranded nucleic acids on polyacrylamide and agarose gels by using glyoxal and acridine orange. Proc Natl Acad Sci USA 74: 4835–4838 (1977).

    Google Scholar 

  33. Moreno J, Chrispeels MJ: A lectin gene encodes the a-amylase inhibitor of the common bean. Proc Natl Acad Sci USA 86: 7885–7889 (1989).

    Google Scholar 

  34. Okamuro JK, Jofuku KD, Goldberg RB: Soybean seed lectin gene and flanking nonseed protein genes are developmentally regulated in transformed tobacco plants. Proc Natl Acad Sci USA 83: 8240–8244 (1986).

    Google Scholar 

  35. Osborn TC, Alexander DC, Sun SSM, Cardona C, Bliss FA: Insecticidal activity and lectin homology of arcelin seed protein. Science 240: 207–210 (1989).

    Google Scholar 

  36. Prasthofer T, Phillips SR, Suddath FL, Engler JA: Design, expression and crystallization of recombinant lectin from the garden pea (Pisum sativum). J Biol Chem 264: 6793–6796 (1989).

    Google Scholar 

  37. Pratt RC, Singh NK, Shade RE, Murdock LL, Bressan RA: Isolation and partial characterisation of a seed lectin from tepary bean that delays bruchid beetle development Plant Physiol 93: 1453–1459 (1990).

    Google Scholar 

  38. Rini JM, Hofmann T, Carver JP: Amino acid sequence differences in the alpha chains of pea seed isolectins: C-terminal processing. Biochem Cell Biol 65: 338–344 (1987).

    Google Scholar 

  39. Rosahl S, Schell J, Willmitzer L: Expression of a tuberspecific storage protein in transgenic potato plants: demonstration of an esterase activity. EMBO J 6: 1155–1159 (1987).

    Google Scholar 

  40. Sengupta-Gopalan C, Reichert NA, Barker RF, Hall TC: Developmentally regulated expression of the bean β phaseolin gene in tobacco seed. Proc Natl Acad Sci USA 82: 3220–3324 (1985).

    Google Scholar 

  41. Shagger H, Von Jagow G: Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range of 1 to 100 kDa. Analyt Biochem 166: 368–379 (1987).

    Google Scholar 

  42. Sharon N, Lis H: Lectins: cell-agglutinating and sugarspecific proteins. Science 177: 949–959 (1972).

    Google Scholar 

  43. Sonnewald U, Sturm A, Chrispeels MJ, Willmitzer L: Targetting and glycosylation of patatin, the major potato tuber protein in leaves of transgenic tobacco. Planta 179: 171–180 (1989).

    Google Scholar 

  44. Sturm A, Voelker TA, Herman EM, Chrispeels MJ: Correct glycosylation, golgi-processing and targeting to protein bodies of the vacuolar protein phytohemaglutinin in transgenic tobacco. Planta 175: 170–183 (1988).

    Google Scholar 

  45. Thomas PS: Hybridization of denatured RNA and DNA fragments transfered to nitrocellulose. Proc Natl Acad Sci USA 77: 5201–5205 (1980).

    Google Scholar 

  46. Trowbridge IS: Isolation and characterization of a mitogenic lectin fromPisum sativum. J Biol Chem 249: 6004–6012 (1974).

    Google Scholar 

  47. Voelker TA, Herman EM, Chrispeels MJ: In vitro mutated phytohemagglutinin genes expressed in tobacco seeds: Role of glycans in protein targetting and protein stability. Plant Cell 1: 95–104 (1989).

    Google Scholar 

  48. Wilkins TA, Bednarek SY, Raikhel NV: Role of propeptide glycan in post-translational processing and transport of barley lectin to vacuoles in transgenic tobacco. Plant Cell 2: 301–313 (1990).

    Google Scholar 

  49. Zambryski P, Joos H, Genetello C, Leemans J, Van Montagu M, Schell J: Ti plasmid vector for the introduction of DNA into plant cells without alteration of their normal regeneration capacity. EMBO J 2: 2143–2150 (1983).

    Google Scholar 

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Author notes

  1. Glyn A. Edwards

    Present address: Shell Research Ltd., Sittingbourne Research Centre, ME9 8AG, Sittingbourne, Kent, UK

Authors and Affiliations

  1. Department of Biological Science, Durham University, DH1 3LE, Durham, Co. Durham, UK

    Glyn A. Edwards & Donald Boulter

  2. Shell Research Ltd., Sittingbourne Research Centre, ME9 8AG, Sittingbourne, Kent, UK

    Andrew Hepher & Stephen P. Clerk

Authors
  1. Glyn A. Edwards
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  2. Andrew Hepher
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  3. Stephen P. Clerk
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  4. Donald Boulter
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Edwards, G.A., Hepher, A., Clerk, S.P. et al. Pea lectin is correctly processed, stable and active in leaves of transgenic potato plants. Plant Mol Biol 17, 89–100 (1991). https://doi.org/10.1007/BF00036809

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  • DOI: https://doi.org/10.1007/BF00036809

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