Characterization and distribution of chymotrypsin-like and other digestive proteases in Colorado potato beetle larvae

Novillo, Concepción; Castañera, Pedro; Ortego, Félix

doi:10.1002/(sici)1520-6327(1997)36:3<181::aid-arch3>3.0.co;2-x

Cited by 75 publications

(47 citation statements)

References 38 publications

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“…During larval development, maximum specific azocaseinase activity at pH 7.5-8.5 was detected in the 5th day of larval life, and zymogram showed the presence of two alkaline proteinases on the 3rd day, with accentuated activity on the 5th day, disappearing on the 6th day, coinciding with the pupae period, and appearing in the adult insect gut. Similar patterns of specific digestive proteinase activity were also reported throughout the larval development of Lepidoptera (Broadway and Duffey, 1985) and in Coleoptera (Michaud et al, 1995;Blanco-Labra et al, 1996;Novillo et al, 1997;Alarcon et al, 2002). In Diptera such as Lucilia cuprina (Casu et al, 1996) and Chrysomya bezziana (Muharsini et al, 2000), two serine proteinases in the larval lumen sensitive to specific inhibitors of trypsin-like enzymes were detected and isolated.…”

Section: Discussionsupporting

confidence: 71%

Digestive enzymes during development of Ceratitis capitata (Diptera:Tephritidae) and effects of SBTI on its digestive serine proteinase targets

Silva

Alcazar

Macedo

et al. 2006

Insect Biochemistry and Molecular Biology

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The digestive system of Ceratitis capitata was characterized during its larval development and in the insect stage. Disaccharidases against maltose and sucrose were more evident in the 2nd and 3rd day of larval development and in the adult stage, respectively. Glycosil-hydrolyses with higher specific a-galactosidasic and b-galactosidasic activities were detected in the 2nd and 3rd day of the larval stage, respectively. Specific proteolytic activities against azocasein showed an increase in the 4th and 5th day of the larval stage and in the adult stage. Specific hemoglobin activities were constant between 2nd and 6th day of the larval stage. The larvae used mainly serine proteinases, such as trypsin/chymotrypsin, and the adult insects only chymotrypsin-like enzymes in their digestive process. Two serine proteinases were separated from zymogram between the 4th and 5th day of larval development and in the adult stage. Effect of soybean trypsin inhibitor (SBTI, a serine proteinase inhibitor) on development of C. capitata was examined by bioassay. C. capitata was susceptible to SBTI which affected larval mass at ED 50 3.01%. r

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Section: Discussionsupporting

confidence: 71%

Digestive enzymes during development of Ceratitis capitata (Diptera:Tephritidae) and effects of SBTI on its digestive serine proteinase targets

Silva

Alcazar

Macedo

et al. 2006

Insect Biochemistry and Molecular Biology

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“…3A) but merely detectable at pH 6.5 (not shown), which points again to the presence of serine proteinases in the midgut extracts, as also reported for other Coleoptera predominantly relying on cysteine proteases for dietary protein digestion (e.g. Novillo et al, 1997;Vinokurov et al, 2006a,b;Prabhakar et al, 2007).…”

Section: Cysteine and Serine Proteases In The Banana Weevil Midgutsupporting

confidence: 77%

Deleterious effects of plant cystatins against the banana weevil Cosmopolites sordidus

Kiggundu

Muchwezi

Vyver

et al. 2009

Arch Insect Biochem Physiol

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The general potential of plant cystatins for the development of insect-resistant transgenic plants still remains to be established given the natural ability of several insects to compensate for the loss of active cysteine proteases following inhibitor ingestion. Here we assessed the potential of cystatins for the development of banana lines resistant to the banana weevil Cosmopolites sordidus, a major pest of banana and plantain in Africa. Protease inhibitory assays were first conducted with protein and methylcoumarin (MCA) peptide substrates to measure the inhibitory efficiency of different cystatins in vitro, followed by a diet bioassay with cystatin-infiltrated banana stem disks to monitor the impact of two plant cystatins, oryzacystatin I (OC-I, or OsCYS1)and papaya cystatin (CpCYS1), on the overall growth rate of young weevil larvae. As observed earlier for other Coleoptera, banana weevils produce a variety of proteases for dietary protein digestion, including in particular Z-Phe-Arg-MCA-hydrolyzing (cathepsin L-like) and Z-Arg-Arg- MCA-hydrolyzing (cathepsin B-like) proteases active in mildly acidic conditions. Both enzyme populations were sensitive to the diagnostic cysteine protease inhibitor E-64 and to different plant cystatins including OsCYS1. In line with these broad inhibitory effects of cystatins, OsCYS1 andCpCYS1 caused an important growth delay in young larvae developing for 10 days in cystatininfiltrated banana stem disks. These promising results, which illustrate the natural susceptibility of C. sordidus to plant cystatins, are discussed in the light of current genomic data on coleopteran cysteine cathepsins and recent hypotheses suggesting a key role for digestive cathepsin B-like enzymes as a determinant for resistance or susceptibility to plant cystatins in Coleoptera.Keywords: Banana weevil (Cosmopolites sordidus); banana (Musa spp.); cathepsin B-like proteases; cysteine cathepsins; plant cystatins; oryzacystatin I Plant cystatins against the banana weevil Page 3 INTRODUCTIONThe banana weevil Cosmopolites sordidus (Germar) (Coleoptera: Curculionidae) is an insect pest of considerable importance in Africa, associated with the rapid decline of banana and plantain plantations (Gold, 1999a(Gold, , 2000Swennen and Vuylsteke, 2001;Gold et al., 2005). At the adult stage, banana weevil females deposit their eggs inside host plant tissues, at the base of the pseudostem or on an exposed corm. On hatching, the larvae tunnel through the corm to feed and develop, where they damage tissues, compromise water and mineral uptake, and weaken the colonized organs. Banana weevil infestations cause important losses in the field by a direct negative impact on harvestable bunch weight and a weakening effect on infested organs causing plant toppling during windstorms (Sengooba, 1986;Rukazambuga et al., 1998).Banana weevil control in Africa relies essentially on cultural and sanitation practices such as the use of clean planting material, the systematic trapping of adult weevils to prevent population build-up, an...

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“…Cathepsin D-like activity was assayed in 0.5 mi of reaction buffer containing 10 ug of mite extract, using 0.2% haemoglobin as described by Novillo et al (1997). Carboxypeptidases A and B-like activities were assayed in 0.5 mi of reaction buffer containing 20 ug of mite extracts, using 1 mM HPA (hippuryl-phenylalanine) for carboxypeptidase A-like activities and 1 mM HA (hippuryl-L-arginine) for carboxypeptidase B-like activities as described by Ortego et al (1996).…”

Section: Gelatin-page Gelsmentioning

confidence: 99%

Expression of a barley cystatin gene in maize enhances resistance against phytophagous mites by altering their cysteine-proteases

et al. 2010

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Phytocystatins are inhibitors of cysteine-proteases from plants putatively involved in plant defence based on their capability of inhibit heterologous enzymes. We have previously characterised the whole cystatin gene family members from barley (HvCPI-1 to HvCPI-13). The aim of this study was to assess the effects of barley cystatins on two phytophagous spider mites, Tetranychus urticae and Brevipalpus chilensis. The determination of proteolytic activity profile in both mite species showed the presence of the cysteine-proteases, putative targets of cystatins, among other enzymatic activities. All barley cystatins, except HvCPI-1 and HvCPI-7, inhibited in vitro mite cathepsin L-and/or cathepsin B-like activities, HvCPI-6 being the strongest inhibitor for both mite species. Transgenic maize plants expressing HvCPI-6 protein were generated and the functional integrity of the cystatin transgene was confirmed by in vitro inhibitory effect observed against T urticae and B. chilensis protein extracts. Feeding experiments impaired on transgenic lines performed with T urticae impaired mite development and reproductive performance. Besides, a significant reduction of cathepsin L-like and/or cathepsin B-like activities was observed when the spider mite fed on maize plants expressing HvCPI-6 cystatin. These findings reveal the potential of barley cystatins as acaricide proteins to protect plants against two important mite pests.

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Characterization and distribution of chymotrypsin-like and other digestive proteases in Colorado potato beetle larvae

Cited by 75 publications

References 38 publications

Digestive enzymes during development of Ceratitis capitata (Diptera:Tephritidae) and effects of SBTI on its digestive serine proteinase targets

Digestive enzymes during development of Ceratitis capitata (Diptera:Tephritidae) and effects of SBTI on its digestive serine proteinase targets

Deleterious effects of plant cystatins against the banana weevil Cosmopolites sordidus

Expression of a barley cystatin gene in maize enhances resistance against phytophagous mites by altering their cysteine-proteases

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