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A cold inducible multidomain cystatin from winter wheat inhibits growth of the snow mold fungus, Microdochium nivale

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Abstract

A novel cold-induced cystatin cDNA clone (TaMDC1) was isolated from cold acclimated winter wheat crown tissue by using a macroarray-based differential screening method. The deduced amino acid sequence consisted of a putative N-terminal secretory signal peptide of 37 amino acids and a mature protein (mTaMDC1) with a molecular mass of 23 kDa. The mTaMDC1 had a highly conserved N-terminal cystatin domain and a long C-terminal extension containing a second region, which exhibited partial similarity to the cystatin domain. The recombinant mTaMDC1 was purified from Escherichia coli and its cysteine proteinase inhibitory activity against papain was analyzed. The calculated Ki value of 5.8×10−7 M is comparable to those reported for other phytocystatins. Northern and western blot analyses showed elevated expression of TaMDC1 mRNA and protein during cold acclimation of wheat. In addition to cold, accumulation of the TaMDC1 message was induced by other abiotic stresses including drought, salt and ABA treatment. Investigation of in vitro antifungal activity of mTaMDC1 showed strong inhibition on the mycelium growth of the snow mold fungus Microdochium nivale. Hyphae growth was totally inhibited in the presence of 50 μg/ml mTaMDC1 and morphological changes such as swelling, fragmentation and sporulation of the fungus were observed. The mechanisms of the in vitro antifungal effects and the possible involvement of TaMDC1 in cold induced snow mold resistance of winter wheat are discussed.

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Abbreviations

WC:

Wheat cystatin

CA:

Cold acclimation

NA:

Non-acclimation

DA:

De-acclimation

GST:

Gluthatione-S-transferase

IPTG:

Isopropyl-β-D-thiogalactoside

BAPA:

Benzoyl-L-arginine-p-nitroanilide

PMSF:

Phenylmethylsulfonyl fluoride

ABA:

Abscisic acid

PDB (A):

Potato dextrose broth (agar)

BSA:

Bovine serum albumin

References

  • Abe K, Emori Y, Kondo H, Suzuki K, Arai S (1987) 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 262:16793–16797

    PubMed  CAS  Google Scholar 

  • Abe K, Kondo H, Watanabe H, Emori Y, Arai S (1991) Oryzacystatins as the first well-defined cystatins of plant origin and their target proteinases in rice seeds. Biomed Biochim Acta 50:637–641

    PubMed  CAS  Google Scholar 

  • Abe M, Abe K, Iwabuchi K, Domoto C, Arai S (1994) Corn cystatin I expressed in Escherichia coli: investigation of its inhibitory profile and occurrence in corn kernels. J Biochem (Tokyo) 116:488–492

    CAS  Google Scholar 

  • Abe M, Arai S (1991) Some properties of a cysteine proteinase inhibitor from corn endosperm. Agric Biol Chem 55:2417–2418

    PubMed  CAS  Google Scholar 

  • Arai S, Watanabe H, Kondo H, Emori Y, Abe K (1991) 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 (Tokyo) 109:294–298

    CAS  Google Scholar 

  • Barret AJ (1987) The cystatins: a new class of peptidase inhibitors. Trends Biochem Sci 12:193–196

    Article  Google Scholar 

  • Brzin J, Popovic T, Ritonja A, Puizdar V, Kidric M (1998) Related cystatin inhibitors from leaf and from seed of Phaseolus vulgaris L. Plant Sci 138:17–26

    Article  CAS  Google Scholar 

  • Cipriani G, Fuentes S, Bello V, Salazar LF, Ghislain M, Zhang DP (2000) Transgene expression of rice cysteine proteinase inhibitor for the development of resistance against sweet potato feathery mottle virus. Program Repport 1999–2000, International popato center, Lima, Peru pp 267–271

  • Corre-Menguy F, Cejudo FJ, Mazubert C, Vidal J, Lelandais-Briere C, Torres G, Rode A, Hartmann C (2002) Characterization of the expression of a wheat cystatin gene during caryopsis development. Plant Mol Biol 50:687–698

    Article  CAS  Google Scholar 

  • Diop NN, Kidric M, Repellin A, Gareil M, d’Arcy-Lameta A, Pham Thi AT, Zuily-Fodil Y (2004) A multicystatin is induced by drought-stress in cowpea (Vigna unguiculata (L.) Walp.) leaves. FEBS Lett 577:545–550

    Article  PubMed  CAS  Google Scholar 

  • Fernandes KVS, Campos FAP, R.R. DV, Xavier-Filho J (1991) Expression of papain inhibitors during development of cowpea seeds. Plant Sci 74:179–184

    Article  CAS  Google Scholar 

  • Gaddour K, Vicente-Carbajosa J, Lara P, Isabel-Lamoneda I, Diaz I, Carbonero P (2001) A constitutive cystatin-encoding gene from barley (Icy) responds differentially to abiotic stimuli. Plant Mol Biol 45:599–608

    Article  PubMed  CAS  Google Scholar 

  • Gaudet DA, Laroche A (1998) Winter survival of cereals parasitized by snow mold. In: Li PH, Chen THH (eds) Plant cold hardiness. Plenum, New York, pp 331–342

    Google Scholar 

  • Guy CL (1990) Cold acclimation and freezing stress tolerance: role of protein methabolism. Annu Rev Plant Physiol Plant Mol Biol 41:187–223

    CAS  Google Scholar 

  • Hines ME, Osuala CI, Nielsen SS (1991) Isolation and partial characterization of a soybean cystatin cysteine proteinase inhibitor of coleopteran digestive proteolytic activity. J Agric Food Chem 39:1515–1520

    Article  CAS  Google Scholar 

  • Hon WC, Griffith M, Mlynarz A, Kwok YC, Yang DS (1995) Antifreeze proteins in winter rye are similar to pathogenesis-related proteins. Plant Physiol 109:879–889

    Article  PubMed  CAS  Google Scholar 

  • Joshi BN, Sainani MN, Bastawade KB, Gupta VS, Ranjekar PK (1998) Cysteine protease inhibitor from pearl millet: a new class of antifungal protein. Biochem Biophys Res Commun 246:382–387

    Article  PubMed  CAS  Google Scholar 

  • Kimura M, Ikeda T, Fukumoto D, Yamasaki N, Yonekura M (1995) Primary structure of a cysteine proteinase inhibitor from the fruit of avocado (Persea americana Mill). Biosci Biotechnol Biochem 59:2328–2329

    Article  PubMed  CAS  Google Scholar 

  • Koike M, Okamoto T, Tsuda S, Imai R (2002) A novel plant defensin-like gene of winter wheat is specifically induced during cold acclimation. Biochem Biophys Res Commun 298:46–53

    Article  PubMed  CAS  Google Scholar 

  • Kondo H, Abe K, Emori Y, Arai S (1991) Gene organization of oryzacystatin-II, a new cystatin superfamily member of plant origin, is closely related to that of oryzacystatin-I but different from those of animal cystatins. FEBS Lett 278:87–90

    Article  PubMed  CAS  Google Scholar 

  • Kondo H, Abe K, Nishimura I, Watanabe H, Emori Y, Arai S (1990) Two distinct cystatin species in rice seeds with different specificities against cysteine proteinases. Molecular cloning, expression, and biochemical studies on oryzacystatin-II. J Biol Chem 265:15832–15837

    PubMed  CAS  Google Scholar 

  • Kouzuma Y, Inanaga H, Doi-Kawano K, Yamasaki N, Kimura M (2000) Molecular cloning and functional expression of cDNA encoding the cysteine proteinase inhibitor with three cystatin domains from sunflower seeds. J Biochem (Tokyo) 128:161–166

    CAS  Google Scholar 

  • Kouzuma Y, Kawano K, Kimura M, Yamasaki N, Kadowaki T, Yamamoto K (1996) Purification, characterization, and sequencing of two cysteine proteinase inhibitors, Sca and Scb, from sunflower (Helianthus annuus) seeds. J Biochem (Tokyo) 119:1106–13

    CAS  Google Scholar 

  • Kuroda M, Kiyosaki T, Matsumoto I, Misaka T, Arai S, Abe K (2001) Molecular cloning, characterization, and expression of wheat cystatins. Biosci Biotechnol Biochem 65:22–8

    Article  PubMed  CAS  Google Scholar 

  • Liang C, Brookhart G, Feng GH, Reeck GR, Kramer KJ (1991) Inhibition of digestive proteinases of stored grain Coleoptera by oryzacystatin, a cysteine proteinase inhibitor from rice seed. FEBS Lett 278:139–142

    Article  PubMed  CAS  Google Scholar 

  • Lim CO, Lee SI, Chung WS, Park SH, Hwang I, Cho MJ (1996) Characterization of a cDNA encoding a cysteine proteinase inhibitor from chinese cabbage (Brassica campestris L. ssp. pekinensis) flower buds. Plant Mol Biol 30:373–379

    Article  PubMed  CAS  Google Scholar 

  • Machida S, Saito M (1993) Purification and characterization of membrane-bound chitin synthase. J Biol Chem 268:1702–1707

    PubMed  CAS  Google Scholar 

  • Marchler-Bauer A, Anderson JB, Cherukuri PF, DeWeese-Scott C, Geer LY, Gwadz M, He S, Hurwitz DI, Jackson JD, Ke Z, Lanczycki CJ, Liebert CA, Liu C, Lu F, Marchler GH, Mullokandov M, Shoemaker BA, Simonyan V, Song JS, Thiessen PA, Yamashita RA, Yin JJ, Zhang D, Bryant SH (2005) CDD: a Conserved Domain Database for protein classification. Nucleic Acids Res 33, Database Issue: D192–196

  • Margis R, Reis EM, Villeret V (1998) Structural and phylogenetic relationships among plant and animal cystatins. Arch Biochem Biophys 359:24–30

    Article  PubMed  CAS  Google Scholar 

  • Martinez M, Lopez-Solanilla E, Rodriguez-Palenzuela P, Carbonero P, Diaz I (2003) Inhibition of plant-pathogenic fungi by the barley cystatin Hv-CPI (gene Icy) is not associated with its cysteine-proteinase inhibitory properties. Mol Plant Microbe Interact 16:876–883

    Article  PubMed  CAS  Google Scholar 

  • Martinez M, Abraham Z, Gambardella M, Echaide M, Carbonero P, Diaz I (2005a) The strawberry gene Cyf1 encodes a phytocystatins with antifungal activity. J Exp Bot 56:1821–1829

    Article  CAS  Google Scholar 

  • Martinez M, Abraham Z, Carbonero P, Diaz I (2005b) Comparative phylogenic analysis of cystatin gene families from Arabidopsis, rice and barley. Mol Gen Genom 273:423–432

    Article  CAS  Google Scholar 

  • Masonneau A, Condamine P, Wisniewski JP, Zivy M, Rogowsky PM (2005) Maize cystatins respond to developmental cues, cold stress and drought. Biochim Biophys Acta 1729:186–199

    Google Scholar 

  • Misaka T, Kuroda M, Iwabuchi K, Abe K, Arai S (1996) Soyacystatin, a novel cysteine proteinase inhibitor in soybean, is distinct in protein structure and gene organization from other cystatins of animal and plant origin. Eur J Biochem 240:609–614

    Article  PubMed  CAS  Google Scholar 

  • Nakajima T, Abe J (1996) Environmental factors effecting expression of resistance to pink snow mold caused by Microdochium nivale in winter wheat. Can J Bot 74:1783–1788

    Article  Google Scholar 

  • Ojima A, Shiota H, Higashi K, Kamada H, Shimma Y, Wada M, Satoh S (1997) An extracellular insoluble inhibitor of cysteine proteinases in cell cultures and seeds of carrot. Plant Mol Biol 34:99–109

    Article  PubMed  CAS  Google Scholar 

  • Oliva ML, Carmona AK, Andrade SS, Cotrin SS, Soares-Costa A, Henrique-Silva F (2004) Inhibitory selectivity of canecystatin: a recombinant cysteine peptidase inhibitor from sugarcane. Biochem Biophys Res Commun 320:1082–1086

    Article  PubMed  CAS  Google Scholar 

  • Pernas M, Lopez-Solanilla E, Sanchez-Monge R, Salcedo G, Rodriguez-Palenzuela P (1999) Antifungal activity of a plant cystatin. Mol Plant Microbe Interact 12:624–627

    Article  CAS  Google Scholar 

  • Pernas M, Sanchez-Monge R, Gomez L, Salcedo G (1998) A chestnut seed cystatin differentially effective against cysteine proteinases from closely related pests. Plant Mol Biol 38:1235–42

    Article  PubMed  CAS  Google Scholar 

  • Pernas M, Sanchez-Monge R, Salcedo G (2000) Biotic and abiotic stress can induce cystatin expression in chestnut. FEBS Lett 467:206–10

    Article  PubMed  CAS  Google Scholar 

  • Ryan SN, McManus MT, Laing WA (2003) Identification and characterisation of proteinase inhibitors and their genes from seeds of apple (Malus domestica). J Biochem (Tokyo) 134:31–42

    CAS  Google Scholar 

  • Salmia MA (1980) Inhibitors of endogenous proteinases in Scots pine seeds: fractionation and activity changes during germination. Physiol Plant 48:266–270

    Article  CAS  Google Scholar 

  • Shyu DJ, Chou W-M, Yiu T-J, Lin CPC, Tzen JTC (2004) Cloning, functional expression, and characterization of cystatin in sesame seed. J Agric Food Chem 52:1350–1356

    Article  PubMed  CAS  Google Scholar 

  • Soares-Costa A, Beltramini LM, Thiemann OH, Henrique-Silva F (2002) A sugarcane cystatin: recombinant expression, purification, and antifungal activity. Biochem Biophys Res Commun 296:1194–1199

    Article  PubMed  CAS  Google Scholar 

  • Sugawara H, Shibuya K, Yoshioka T, Hashiba T, Satoh S (2000) Is a cysteine proteinase inhibitor involved in the regulation of petal wilting in senescing carnation (Dianthus caryophyllus L.) flowers? J Exp Bot 53:407–413

    Article  Google Scholar 

  • Sugawara H, Yoshioka T, Hashiba T, Satoh S (2002) Antifungal activity of a recombinant carnation cystatin, rDC-CPIn. Plant Biotechnol 19:207–209

    CAS  Google Scholar 

  • Tronsmo AM (1993) Resistance to winter stress factors in half-sib families of Dactylis glomerata, tested in a controlled environment. Acta Agric Scand 43:89–96

    Google Scholar 

  • Urwin PE, Lilley CJ, McPherson MJ, Atkinson HJ (1997) Resistance to both cyst and root-knot nematodes conferred by transgenic Arabidopsis expressing a modified plant cystatin. Plant J 12:455–461

    Article  PubMed  CAS  Google Scholar 

  • Waldron C, Wegrich LM, Merlo PA, Walsh TA (1993) Characterization of a genomic sequence coding for potato multicystatin, an eight-domain cysteine proteinase inhibitor. Plant Mol Biol 23:801–12

    Article  PubMed  CAS  Google Scholar 

  • Womack JS, Randall J, Kemp JD (2000) Identification of a signal peptide for oryzacystatin-I. Planta 210:844–847

    Article  PubMed  CAS  Google Scholar 

  • Wu J, Haard NF (2004) Purification and characterization of a cystatin form the leaves of methyl jasmonate-treated tomato plants. Comp Biochem Physiol C Toxicol Pharmacol 127:209–220

    Google Scholar 

  • Yamada T, Ohta H, Shinohara A, Iwamatsu A, Shimada H, Tsuchiya T, Masuda T, Takamiya K (2000) A cysteine protease from maize isolated in a complex with cystatin. Plant Cell Physiol 41:185–91

    PubMed  CAS  Google Scholar 

  • Yang AH, Yeh KW (2005) Molecular cloning, recombinant gene expression, and antifungal activity of cystatin from taro (Colocasia esculenta cv. Kaosiung no.1). Planta 221:493–501

    Article  PubMed  CAS  Google Scholar 

  • Yoza K, Nakamura S, Yaguchi M, Haraguchi K, Ohtsubo K (2002) Molecular cloning and functional expression of cDNA encoding a cysteine proteinase inhibitor, cystatin, from Job’s tears (Coix lacryma-jobi L. var. Ma-yuen Stapf). Biosci Biotechnol Biochem 66:2287–91

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

We would like to thank Dr. Fumihiro Terami (National Agricultural Research Center for Hokkaido Region, Japan) for kindly providing the snow mold strain and comments. This work was supported by a grant from MAFF (BioDesign 1207) to R.I. It was also supported by a JSPS postdoctral felloship to N.K.C.

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Authors and Affiliations

  1. Winter Stress Laboratory, National Agricultural Research Center for Hokkaido Region, Hitsujigaoka 1, 062-8555, Toyohira-ku, Sapporo, Japan

    Petya Koeva Christova, Nikolai Kirilov Christov & Ryozo Imai

  2. AgroBioInstitute, Dragan Tsankov 8, 1164, Sofia, Bulgaria

    Nikolai Kirilov Christov

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  1. Petya Koeva Christova
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  2. Nikolai Kirilov Christov
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  3. Ryozo Imai
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Correspondence to Ryozo Imai.

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Christova, P.K., Christov, N.K. & Imai, R. A cold inducible multidomain cystatin from winter wheat inhibits growth of the snow mold fungus, Microdochium nivale . Planta 223, 1207–1218 (2006). https://doi.org/10.1007/s00425-005-0169-9

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