Antifungal Activity of a Recombinant Carnation Cystatin, rDC-CPIn.

Sugawara, H.; Yoshioka, T.; Hashiba, Teruyoshi; Satoh, Shigeru

doi:10.5511/plantbiotechnology.19.207

Cited by 8 publications

(4 citation statements)

References 24 publications

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“…Cystatins from soybean, rice and chestnut demonstrated antifeedant activity against insects (Hines et al 1991;Liang et al 1991;Pernas et al 1998). In addition, antifungal activity of cystatins from pearl millet, chestnut, sugarcane, carnation, barley, strawberry and taro have been reported in the last few years (Joshi et al 1998;Pernas et al 1999;Soares-Costa et al 2002;Sugawara et al 2002;Martinez et al 2003, 2005a, Yang and Yeh 2005. Here, we described for the first time the antifungal activity of a cystatin to the phytopathogenic fungus M. nivale.…”

Section: Discussionmentioning

confidence: 81%

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

Christova

Christov

Imai

2005

Planta

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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 x 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 mug/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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Section: Discussionmentioning

confidence: 81%

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

Christova

Christov

Imai

2005

Planta

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“…Expression of cystatins at various phases of plant development and in response to stresses has previously been described (Abraham et al 2006, Diop et al 2004, Masonneau et al 2005, Pernas et al 2000). Two possible roles have been proposed: regulation of proteolysis during seed maturation and germination and plant defense by inhibiting exogenous proteinases from pests (Sugawara et al 2002b). In addition, in maize another role, stabilizer of the CPPIC, was reported (Yamada et al 2001b).…”

Section: Discussionmentioning

confidence: 99%

Biochemical and molecular characterization of senescence‐related cysteine protease–cystatin complex from spinach leaf

Tajima

Yamaguchi

Matsushima

et al. 2010

Physiologia Plantarum

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Cysteine proteases (CPs) with N-succinyl-Leu-Tyr-4-methylcoumaryl-7-amide (Suc-LY-MCA) cleavage activity were investigated in green and senescent leaves of spinach. The enzyme activity was separated into two major and several faint minor peaks by hydrophobic chromatography. These peaks were conventionally designated as CP1, CP2 and CP3, according to their order of elution. From the analyses of molecular mass, subunit structure, amino acid sequences and cDNA cloning, CP2 was a monomer complex (SoCP-CPI) (51 kDa) composed of a 41-kDa core protein, SoCP (Spinacia oleracea cysteine protease), and 14-kDa cystatin, a cysteine protease inhibitor (CPI), while CP3 was a trimer complex (SoCP-CPI)(3) (151 kDa) of the same subunits as SoCP-CPI and showed a wider range of specificity toward natural substrates than SoCP-CPI. Trimer (SoCP-CPI)(3) was irreversibly formed from monomers through association. The results of reverse transcription-polymerase chain reaction (RT-PCR) revealed that mRNAs of CPI and SoCP are hardly expressed in green leaves, but they are coordinately expressed in senescent leaves, suggesting that these proteases involve in senescence. Purified recombinant CPI had strong inhibitory activity against trimer SoCP, (SoCP)(3) , which had a cystatin deleted with K(i) value of 1.33 × 10(-9) M. After treatment of the enzyme with a succinate buffer (pH 5) at the most active pH of the enzyme, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and activity analyses showed that cystatin was released from both monomer SoCP-CPI and trimer (SoCP-CPI)(3) complexes with a concomitant activation. Thus, the removal of a cystatin is necessary to activate the enzyme activity.

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“…They have been related with defence mechanisms against insects and plant pathogens as well as plant response to heat, cold and saline stresses (Pernas et al 2000; Carrillo et al 2011; Valdés-Rodríguez et al 2015). It is well documented that phytocystatins present antifungal activity against a wide range of pathogenic fungi such as Botritys cinerea, Plectosphaerella cucumerina, Colletotrichum graminicola, Colletotrichum capsisi, Pyricularia grisea, Fusarium oxysporum, Septoria nodorum, Phytophthora nicotianae, Sclerotiana sclerotiorum, Sclerotium rolfsii, Alternaria brassicae, Glomerella cingulata, Pythium aphanidermatum, Rhizoctonia solani, Trichoderma reesei, Aspergillus sydowii, Helminthosporium sesamum (Pernas et al 1999; Sugawara et al 2002; Martínez et al 2003; Yang & Yeh 2005; Abraham et al 2006; Porruan et al 2013; Cheng et al 2014). However, the mechanism by which phytocystatins inhibit fungal growth is not yet known.…”

Section: Introductionmentioning

confidence: 99%

In vitroeffect of recombinant amaranth cystatin (AhCPI) on spore germination, mycelial growth, stress response and cellular integrity ofAspergillus nigerandAspergillus parasiticus

et al. 2015

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The inhibitory effect of recombinant amaranth cystatin (AhCPI) on the spore germination and growth of the mycotoxigenic fungus Aspergillus parasiticus and Aspergillus niger was investigated. AhCPI showed a concentration-dependent antifungal activity against both fungi. Differential effects were observed when fungi were treated with cystatin in two developmental stages. When AhCPI was added to young mycelium cultures of A. niger, it had a dramatic effect on mycelial growth compared with old mycelium cultures. On the contrary, there was no differential effect of AhCPI addition to either old or young mycelium of A. parasiticus. Furthermore, electron microscopic observations showed that cystatin caused important effects at the level of cell morphology and organelle integrity of both fungi. Additionally, A. parasiticus spores treated with AhCPI presented sensitivity to oxidative, osmotic and ionic stresses; in opposition, under same conditions, A. niger did not show sensitivity to any stressful agent. These results suggest that AhCPI antifungal activity might be related with damage to cell integrity, affecting the survival of the fungi. In addition, our evidences showed that fungal species respond dissimilarly to cystatin; however, such disparities can be used to the control of unwanted fungi.

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Antifungal Activity of a Recombinant Carnation Cystatin, rDC-CPIn.

Cited by 8 publications

References 24 publications

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

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

Biochemical and molecular characterization of senescence‐related cysteine protease–cystatin complex from spinach leaf

In vitroeffect of recombinant amaranth cystatin (AhCPI) on spore germination, mycelial growth, stress response and cellular integrity ofAspergillus nigerandAspergillus parasiticus

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