Skip to main content
SpringerLink
Log in

Isolation and characterization of a gene encoding endo-β-1,4-glucanase from pepper (Capsicum annuum L.)

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

The endo-β-1,4-glucanases, or cellulases, of higher plants are cell wall-associated enzymes believed to function in cell wall changes associated with the diverse processes of fruit ripening, organ abscission and cell elongation. We have isolated and characterized cDNA and genomic clones encoding a cellulase, PCEL1, which is abundant in ripening pepper fruit. Genomic analysis indicates that PCEL1 is encoded by a single gene, PCEL1, which belongs to a small, structurally divergent gene family. In ripening fruit, PCEL1 transcription is initiated at two distinct sites which yields overlapping mRNA species of 1.7 and 2.1 kb. High-level accumulation of both transcripts occurs in red fruit, while the 1.7 kb transcript is detected at a much lower level in stem and petiolar tissue. The increase in cellulase activity which is measured during fruit ripening is the product of PCEL1 expression and is tightly coupled to fruit reddening. High-level applications of ethylene serve to enhance the rate of ripening and the accumulation of PCEL1 mRNA. A direct role for ethylene in regulating PCEL1 expression is shown by the exclusive induction, in immature green fruit, of the 1.7 kb transcript in response to prolonged high-level exposure to ethylene - a pattern of expression not observed in fruit development on the vine.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Acebes JL, Zarra I: Cell wall glycanases and their activity against the hemicelluloses from pine hypocotyls. Physiol Plant 86: 433–438 (1992).

    Google Scholar 

  2. Awad M, Lewis L: Avocado cellulase: extraction and purification. J Food Sci 45: 1625–1628 (1980).

    Google Scholar 

  3. Beaudry R, Kays S: Effect of ethylene source on abscission of pepper plant organs. HortScience 23: 742–744 (1988).

    Google Scholar 

  4. Bennett A, Christoffersen R: Synthesis and processing of cel-lulase from ripening avocado fruit. Plant Physiol 81: 830–835 (1986).

    Google Scholar 

  5. Bonghi C, Rascio N, Ramina A, Casadoro G: Cellulase and polygalacturonase involvement in the abscission of leaf and fruit explants of peach. Plant Mol Biol 20: 839–848 (1992).

    Google Scholar 

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

    Google Scholar 

  7. Brady C: Fruit ripening. Annu Rev Plant Physiol 38: 155–178 (1987).

    Google Scholar 

  8. Brummell D, Lashbrook C, Bennett A: Plant endo-1,4-_-Dglucanases. In: Himmel M, Baker J, Overend R (eds) Enzymatic Conversion of Biomass for Fuels Production, pp. 100–129. American Chemical Society, Washington, DC (1994).

    Google Scholar 

  9. Cass L, Kirven K, Christoffersen R: Isolation and characterization of a cellulase gene family member expressed during avocado fruit ripening. Mol Gen Genet 223: 76–86 (1990).

    Google Scholar 

  10. Chen J, Varner J: An extracellular matrix protein in plants: characterization of a genomic clone for carot extensin. EMBO J 4: 2145–2151 (1985).

    Google Scholar 

  11. Conrad RS, Sundstrom FU: Calcium and ethephon effects on tabasco pepper leaf and fruit retention and fruit color development. J Am Soc Hort Sci 112: 424–426 (1987).

    Google Scholar 

  12. del Campillo E, Lewis L: Occurence of 9.5 cellulase and other hydrolases in flower reproductive organs undergoing major cell wall disruption. Plant Physiol 99: 1015–1020 (1992).

    Google Scholar 

  13. del Campillo E, Reid P, Sexton R, Lewis L: Occurrence and localization of 9.5 cellulase in abscising and non abscising tissues. Plant Cell 2: 245–254 (1990).

    Google Scholar 

  14. Dunsmuir P, Bond D, Lee K, Gidoni D, Townsend J: The expression of introduced genes in regenerated plants. In: Gelvin S, Schilperoort R (eds) Plant Molecular Biology Manual, pp. 45–59. Kluwer Academic Publishers, Dordrecht, Netherlands (1987).

    Google Scholar 

  15. Durbin M, Lewis L: Cellulases in Phaseolus vulgaris. Meth Enzymol 160: 342–351 (1988).

    Google Scholar 

  16. Ferrarese L, Trainotti L, Moretto P, de Lauretto P, Rascio N, Casadoro G: Differential ethylene-inducible expression of cellulase in pepper plants. Plant Mol Biol 29: 735–747 (1995).

    Google Scholar 

  17. Fischer R, Bennett A: Role of cell wall hydrolases in fruit ripening. Annu Rev Plant Physiol Plant Mol Biol 42: 675–703 (1991).

    Google Scholar 

  18. Fry S: Cellulases, hemicelluloses and auxin-stimulated growth: a possible relationship. Physiol Plant 75: 523–536 (1989).

    Google Scholar 

  19. Gross K, Watada A, Kang M, Kim S, Kim K, Lee S: Biochemical changes associated with the ripening of hot pepper fruit. Physiol Plant 66: 31–36 (1986).

    Google Scholar 

  20. Harpster M, Howie W, Dunsmuir P: Characterization of a PCR fragment encoding 1-aminocyclopropane-1-carboxylate synthase in pepper (Capsicum annuum). J Plant Physiol 147: 661–664 (1996).

    Google Scholar 

  21. Harpster M, Townsend JA, Jones JGJ, Bedbrook J, Dunsmuir P: Relative strengths of the 35S cauliflower mosaic virus, 1′, 2′ and nopaline synthase promoter in tobacco, sugarbeet and oilseed rape callus tissue. Mol Gen Genet 212: 182–190 (1988).

    Google Scholar 

  22. Hatfield R, Nevins D: Characterization of the hydrolytic activity of avocado cellulase. Plant Cell Physiol 27: 541–552 (1986).

    Google Scholar 

  23. Hayashi T, Whon Y, Maclachlan G: Pea xyloglucan and cellulose. II. Hydrolysis by pea end-1,4-_-glucanases. Plant Physiol 75: 605–610 (1984).

    Google Scholar 

  24. Holdsworth M, Laties G: Site-specific binding of a nuclear factor to the carrot extensin gene is influenced by both ethylene and wounding. Planta 179: 17–23 (1989).

    Google Scholar 

  25. Huberman M, Pressman E, Jaffe M: Pith autolysis in plants. IV. The activity of polygalacturonase and cellulase during drought stress induced by autolysis. Plant Cell Physiol 34: 795–801 (1993).

    Google Scholar 

  26. Jen J, Robinson M: Pectolytic enzymes in sweet bell peppers (Capsicum annuum L.). J Food Sci 49: 1085–1087 (1984).

    Google Scholar 

  27. Kemmerer E, Tucker M: Comparative study of cellulases associated with adventitious root initiation, apical buds, and leaf, flower, and pod abscission zones in soybean. Plant Physiol 104: 557–562 (1994).

    Google Scholar 

  28. Laemmli U: Cleavage of structural proteins during the assembly of the phage head of bacteriophage T4. Nature 227: 680–685 (1970).

    Google Scholar 

  29. Lashbrook C, Gonzalez-Bosch C, Bennett A: Two divergent endo-β-1,4-glucanase genes exhibit overlapping expression in ripening fruit and abscising flowers. Plant Cell 6: 1485–149 (1994).

    Google Scholar 

  30. Lund P, Lee R, Dunsmuir P: A plant signal sequence enhances the secretion of bacterial ChiA in transgenic tobacco. Plant Physiol 88: 47–53 (1989).

    Google Scholar 

  31. Maclachlan G, Brady C: Multiple forms of 1,4-glucanase in ripening tomato fruits include a xyloglucanase activatable by xyloglucan oligosaccharides. Aust J Plant Physiol 19: 137–146 (1992).

    Google Scholar 

  32. Melton DA, Kreig PA, Rebagliati MR, Manaitis T, Zinn K, Green MR: Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucl Acids Res 12: 7035–7056 (1984).

    Google Scholar 

  33. Mueller G, McKeown R, Corotto L, Hague C, Warren G: Inhibition of recrystallization in ice by chimeric proteins containing antifreeze domains. J Biol Chem 266: 7339–7344 (1991).

    Google Scholar 

  34. Sanger F, Nicklen S, Coulsen A: DNA sequencing with chain terminating inhibitors. Proc Natl Acad Sci USA 74: 5463–5467 (1977).

    Google Scholar 

  35. Schibler U, Sierra F: Alternate promoters in developmental gene expression. Annu Rev Genet 21: 237–257 (1987).

    Google Scholar 

  36. Sexton R, Durbin M, Lewis L, Thomson W: Use of cellulase antibodies to study leaf abscission. Nature 283: 8743–874 (1980).

    Google Scholar 

  37. Taylor J, Coupe S, Picton S, Roberts J: Characterization and accumulation pattern of an mRNA encoding an abscission-related β-1,4-glucanase from leaflets of Sambucus nigra. Plant Mol Biol 24: 961–964 (1994).

    Google Scholar 

  38. Tucker G, Schindler C, Roberts J: Flower abscission in mutant tomato plants. Planta 160: 164–167 (1984).

    Google Scholar 

  39. Tucker M, Sexton R, Del Campillo E, Lewis L: Bean abscission cellulase. Plant Physiol 88: 1257–1262 (1988).

    Google Scholar 

  40. Verma D, Maclachlan G, Byrne H, Ewings D: Regulation and in vitro translation of messenger ribonucleic acid for cellulase from auxin-treated pea epicotyls. J Biol Chem 250: 1019–1026 (1975).

    Google Scholar 

  41. von Heijne G: A new method for predicting signal sequence cleavage sites. Nucl Acids Res 14: 4683–4690 (1986).

    Google Scholar 

  42. Wu S-C, Blumer J, Darvill A, Albersheim P: Characterization of an endo-β-1,4-glucanase gene induced by auxin in elongating pea epicotyls. Plant Physiol 110: 163–170 (1996).

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. DNA Plant Technology Corporation, 6701 San Pablo Avenue, Oakland, CA, 94608, USA

    Mark H. Harpster, Kathleen Y. Lee & Pamela Dunsmuir

Authors
  1. Mark H. Harpster
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Kathleen Y. Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Pamela Dunsmuir
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Harpster, M.H., Lee, K.Y. & Dunsmuir, P. Isolation and characterization of a gene encoding endo-β-1,4-glucanase from pepper (Capsicum annuum L.). Plant Mol Biol 33, 47–59 (1997). https://doi.org/10.1023/A:1005795028489

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1005795028489

Search

Navigation