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. 1996 Aug;111(4):1299–1306. doi: 10.1104/pp.111.4.1299

Two wound-inducible soybean cysteine proteinase inhibitors have greater insect digestive proteinase inhibitory activities than a constitutive homolog.

Y Zhao 1, M A Botella 1, L Subramanian 1, X Niu 1, S S Nielsen 1, R A Bressan 1, P M Hasegawa 1
PMCID: PMC161012  PMID: 8756506

Abstract

Diverse functions for three soybean (Glycine max L. Merr.) cysteine proteinase inhibitors (CysPIs) are inferred from unique characteristics of differential regulation of gene expression and inhibitory activities against specific Cys proteinases. Based on northern blot analyses, we found that the expression in leaves of one soybean CysPI gene (L1) was constitutive and the other two (N2 and R1) were induced by wounding or methyl jasmonate treatment. Induction of N2 and R1 transcript levels in leaves occurred coincidentally with increased papain inhibitory activity. Analyses of kinetic data from bacterial recombinant CysPI proteins indicated that soybean CysPIs are noncompetitive inhibitors of papain. The inhibition constants against papain of the CysPIs encoded by the wound and methyl jasmonate-inducible genes (57 and 21 nM for N2 and R1, respectively) were 500 to 1000 times lower than the inhibition constant of L1 (19,000 nM). N2 and R1 had substantially greater inhibitory activities than L1 against gut cysteine proteinases of the third-instar larvae of western corn rootworm and Colorado potato beetle. Cysteine proteinases were the predominant digestive proteolytic enzymes in the guts of these insects at this developmental stage. N2 and R1 were more inhibitory than the epoxide trans-epoxysuccinyl-L-leucylamide-(4-guanidino)butane (E-64) against western corn rootworm gut proteinases (50% inhibition concentration = 50, 200, and 7000 nM for N2, R1, and E-64, respectively). However, N2 and R1 were less effective than E-64 against the gut proteinases of Colorado potato beetle. These results indicate that the wound-inducible soybean CysPIs, N2 and R1, function in host plant defense against insect predation, and that substantial variation in CysPI activity against insect digestive proteinases exists among plant CysPI proteins.

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Selected References

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  1. Abe K., Emori Y., Kondo H., Arai S., Suzuki K. The NH2-terminal 21 amino acid residues are not essential for the papain-inhibitory activity of oryzacystatin, a member of the cystatin superfamily. Expression of oryzacystatin cDNA and its truncated fragments in Escherichia coli. J Biol Chem. 1988 Jun 5;263(16):7655–7659. [PubMed] [Google Scholar]
  2. Abe K., Emori Y., Kondo H., Suzuki K., Arai S. Molecular cloning of a cysteine proteinase inhibitor of rice (oryzacystatin). Homology with animal cystatins and transient expression in the ripening process of rice seeds. J Biol Chem. 1987 Dec 15;262(35):16793–16797. [PubMed] [Google Scholar]
  3. Abe M., Abe K., Iwabuchi K., Domoto C., Arai S. Corn cystatin I expressed in Escherichia coli: investigation of its inhibitory profile and occurrence in corn kernels. J Biochem. 1994 Sep;116(3):488–492. doi: 10.1093/oxfordjournals.jbchem.a124551. [DOI] [PubMed] [Google Scholar]
  4. Abe M., Abe K., Kuroda M., Arai S. Corn kernel cysteine proteinase inhibitor as a novel cystatin superfamily member of plant origin. Molecular cloning and expression studies. Eur J Biochem. 1992 Nov 1;209(3):933–937. doi: 10.1111/j.1432-1033.1992.tb17365.x. [DOI] [PubMed] [Google Scholar]
  5. Arai S., Watanabe H., Kondo H., Emori Y., Abe K. Papain-inhibitory activity of oryzacystatin, a rice seed cysteine proteinase inhibitor, depends on the central Gln-Val-Val-Ala-Gly region conserved among cystatin superfamily members. J Biochem. 1991 Feb;109(2):294–298. [PubMed] [Google Scholar]
  6. Baumgartner B., Chrispeels M. J. Partial characterization of a protease inhibitor which inhibits the major endopeptidase present in the cotyledons of mung beans. Plant Physiol. 1976 Jul;58(1):1–6. doi: 10.1104/pp.58.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bode W., Engh R., Musil D., Thiele U., Huber R., Karshikov A., Brzin J., Kos J., Turk V. The 2.0 A X-ray crystal structure of chicken egg white cystatin and its possible mode of interaction with cysteine proteinases. EMBO J. 1988 Aug;7(8):2593–2599. doi: 10.1002/j.1460-2075.1988.tb03109.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Botella M. A., Quesada M. A., Medina M. I., Pliego F., Valpuesta V. Induction of a tomato peroxidase gene in vascular tissue. FEBS Lett. 1994 Jun 27;347(2-3):195–198. doi: 10.1016/0014-5793(94)00542-7. [DOI] [PubMed] [Google Scholar]
  9. Colella R., Sakaguchi Y., Nagase H., Bird J. W. Chicken egg white cystatin. Molecular cloning, nucleotide sequence, and tissue distribution. J Biol Chem. 1989 Oct 15;264(29):17164–17169. [PubMed] [Google Scholar]
  10. Farmer E. E., Ryan C. A. Octadecanoid Precursors of Jasmonic Acid Activate the Synthesis of Wound-Inducible Proteinase Inhibitors. Plant Cell. 1992 Feb;4(2):129–134. doi: 10.1105/tpc.4.2.129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fernandes K. V., Sabelli P. A., Barratt D. H., Richardson M., Xavier-Filho J., Shewry P. R. The resistance of cowpea seeds to bruchid beetles is not related to levels of cysteine proteinase inhibitors. Plant Mol Biol. 1993 Oct;23(1):215–219. doi: 10.1007/BF00021433. [DOI] [PubMed] [Google Scholar]
  12. Goetting-Minesky M. P., Mullin B. C. Differential gene expression in an actinorhizal symbiosis: evidence for a nodule-specific cysteine proteinase. Proc Natl Acad Sci U S A. 1994 Oct 11;91(21):9891–9895. doi: 10.1073/pnas.91.21.9891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Guan K. L., Dixon J. E. Eukaryotic proteins expressed in Escherichia coli: an improved thrombin cleavage and purification procedure of fusion proteins with glutathione S-transferase. Anal Biochem. 1991 Feb 1;192(2):262–267. doi: 10.1016/0003-2697(91)90534-z. [DOI] [PubMed] [Google Scholar]
  14. Hengartner M. O., Horvitz H. R. Programmed cell death in Caenorhabditis elegans. Curr Opin Genet Dev. 1994 Aug;4(4):581–586. doi: 10.1016/0959-437x(94)90076-f. [DOI] [PubMed] [Google Scholar]
  15. Hildmann T., Ebneth M., Peña-Cortés H., Sánchez-Serrano J. J., Willmitzer L., Prat S. General roles of abscisic and jasmonic acids in gene activation as a result of mechanical wounding. Plant Cell. 1992 Sep;4(9):1157–1170. doi: 10.1105/tpc.4.9.1157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Holwerda B. C., Rogers J. C. Purification and characterization of aleurain : a plant thiol protease functionally homologous to Mammalian cathepsin h. Plant Physiol. 1992 Jul;99(3):848–855. doi: 10.1104/pp.99.3.848. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jongsma M. A., Bakker P. L., Peters J., Bosch D., Stiekema W. J. Adaptation of Spodoptera exigua larvae to plant proteinase inhibitors by induction of gut proteinase activity insensitive to inhibition. Proc Natl Acad Sci U S A. 1995 Aug 15;92(17):8041–8045. doi: 10.1073/pnas.92.17.8041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kalinski A., Melroy D. L., Dwivedi R. S., Herman E. M. A soybean vacuolar protein (P34) related to thiol proteases is synthesized as a glycoprotein precursor during seed maturation. J Biol Chem. 1992 Jun 15;267(17):12068–12076. [PubMed] [Google Scholar]
  19. Koizumi M., Yamaguchi-Shinozaki K., Tsuji H., Shinozaki K. Structure and expression of two genes that encode distinct drought-inducible cysteine proteinases in Arabidopsis thaliana. Gene. 1993 Jul 30;129(2):175–182. doi: 10.1016/0378-1119(93)90266-6. [DOI] [PubMed] [Google Scholar]
  20. Liang C., Brookhart G., Feng G. H., Reeck G. R., Kramer K. J. Inhibition of digestive proteinases of stored grain Coleoptera by oryzacystatin, a cysteine proteinase inhibitor from rice seed. FEBS Lett. 1991 Jan 28;278(2):139–142. doi: 10.1016/0014-5793(91)80102-9. [DOI] [PubMed] [Google Scholar]
  21. Ritonja A., Kopitar M., Jerala R., Turk V. Primary structure of a new cysteine proteinase inhibitor from pig leucocytes. FEBS Lett. 1989 Sep 25;255(2):211–214. doi: 10.1016/0014-5793(89)81093-2. [DOI] [PubMed] [Google Scholar]
  22. Schaffer M. A., Fischer R. L. Analysis of mRNAs that Accumulate in Response to Low Temperature Identifies a Thiol Protease Gene in Tomato. Plant Physiol. 1988 Jun;87(2):431–436. doi: 10.1104/pp.87.2.431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Shapira R., Nuss D. L. Gene expression by a hypovirulence-associated virus of the chestnut blight fungus involves two papain-like protease activities. Essential residues and cleavage site requirements for p48 autoproteolysis. J Biol Chem. 1991 Oct 15;266(29):19419–19425. [PubMed] [Google Scholar]
  24. Thompson C. B. Apoptosis in the pathogenesis and treatment of disease. Science. 1995 Mar 10;267(5203):1456–1462. doi: 10.1126/science.7878464. [DOI] [PubMed] [Google Scholar]
  25. Turk V., Bode W. The cystatins: protein inhibitors of cysteine proteinases. FEBS Lett. 1991 Jul 22;285(2):213–219. doi: 10.1016/0014-5793(91)80804-c. [DOI] [PubMed] [Google Scholar]
  26. Urwin P. E., Atkinson H. J., Waller D. A., McPherson M. J. Engineered oryzacystatin-I expressed in transgenic hairy roots confers resistance to Globodera pallida. Plant J. 1995 Jul;8(1):121–131. doi: 10.1046/j.1365-313x.1995.08010121.x. [DOI] [PubMed] [Google Scholar]
  27. Waldron C., Wegrich L. M., Merlo P. A., Walsh T. A. Characterization of a genomic sequence coding for potato multicystatin, an eight-domain cysteine proteinase inhibitor. Plant Mol Biol. 1993 Nov;23(4):801–812. doi: 10.1007/BF00021535. [DOI] [PubMed] [Google Scholar]

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