Effects of rice cystatin I expression in transgenic potato on Colorado potato beetle larvae

Lecardonnel, Anne; Chauvin, Laura; Jouanin, Lise; Beaujean, Antony; Prévost, Geneviève; Sangwan‐Norreel, B. S.

doi:10.1016/s0168-9452(98)00197-6

Cited by 91 publications

(53 citation statements)

References 39 publications

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“…Benchekroun et al, 1995;Leplé et al 1995;Girard et al, 1998aGirard et al, , 1998bCloutier et al, 1999Cloutier et al, , 2000Lecardonnel et al, 1999;Outchkourov et al, 2004;Ribeiro et al, 2006;Alvarez-Alfageme et al, 2007), but nonconclusive results were obtained in several cases, correlated with the onset of effective adaptive processes allowing target pests to overcome the deleterious effects of digestive protease inhibition. In practice, the inhibitory potency of protease inhibitors is critical for an effective inhibition of target pest digestive proteases in vivo (Urwin et al, 1995), as it directly determines the amount of recombinant protein needed in the modified host plant to provide sufficient antidigestive or pesticidal effects (Jongsma and Bolter, 1997).…”

Section: Discussionmentioning

confidence: 99%

Tailoring the Specificity of a Plant Cystatin toward Herbivorous Insect Digestive Cysteine Proteases by Single Mutations at Positively Selected Amino Acid Sites

et al. 2008

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Plant cystatins, similar to other defense proteins, include hypervariable, positively selected amino acid sites presumably impacting their biological activity. Using 29 single mutants of the eighth domain of tomato (Solanum lycopersicum) multicystatin, SlCYS8, we assessed here the potential of site-directed mutagenesis at positively selected amino acid sites to generate cystatin variants with improved inhibitory potency and specificity toward herbivorous insect digestive cysteine (Cys) proteases. Compared to SlCYS8, several mutants (22 out of 29) exhibited either improved or lowered potency against different model Cys proteases, strongly suggesting the potential of positively selected amino acids as target sites to modulate the inhibitory specificity of the cystatin toward Cys proteases of agronomic significance. Accordingly, mutations at positively selected sites strongly influenced the inhibitory potency of SlCYS8 against digestive Cys proteases of the insect herbivore Colorado potato beetle (Leptinotarsa decemlineata). In particular, several variants exhibited improved potency against both cystatin-sensitive and cystatininsensitive digestive Cys proteases of this insect. Of these, some variants also showed weaker activity against leaf Cys proteases of the host plant (potato [Solanum tuberosum]) and against a major digestive Cys protease of the two-spotted stinkbug Perillus bioculatus, an insect predator of Colorado potato beetle showing potential for biological control. Overall, these observations suggest the usefulness of site-directed mutagenesis at positively selected amino acid sites for the engineering of recombinant cystatins with both improved inhibitory potency toward the digestive proteases of target herbivores and weaker potency against nontarget Cys proteases in the host plant or the environment.

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Section: Discussionmentioning

confidence: 99%

Tailoring the Specificity of a Plant Cystatin toward Herbivorous Insect Digestive Cysteine Proteases by Single Mutations at Positively Selected Amino Acid Sites

et al. 2008

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“…In this sense, Ser PINs are effective against Lepidoptera (Hilder et al, 1987;Johnson et al, 1989;McManus et al, 1994;Duan et al, 1996;Li et al, 1998;De Leo et al, 2001). On the other hand, aspartic and Cys PINs have been used to generate resistance against coleopteran species (Orr et al, 1994;Leplé et al, 1995;Girard et al, 1998;Lecardonnel et al, 1999).…”

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confidence: 99%

A Novel Function for the Cathepsin D Inhibitor in Tomato

2006

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Proteinaceous aspartic proteinase inhibitors are rare in nature and are described in only a few plant species. One of them corresponds to a family of cathepsin D inhibitors (CDIs) described in potato (Solanum tuberosum), involving up to 15 isoforms with a high sequence similarity. In this work, we describe a tomato (Solanum lycopersicum) wound-inducible protein called jasmonic-induced protein 21 (JIP21). Sequence analysis of its cDNA predicted a putative function as a CDI. The JIP21 gene, whose protein has been demonstrated to be glycosylated, is constitutively expressed in flowers, stem, and fruit, and is inducible to high levels by wounding and methyl jasmonate in leaves of tomato plants. The genomic sequence of JIP21 shows that the gene is intronless and reveals the presence of both a methyl jasmonate box (TGACT) and a G-box (CACGT) in the promoter. In contrast to the presumed role of JIP21 based on sequence analysis, a detailed biochemical characterization of the purified protein uncovers a different function as a strong chymotrypsin inhibitor, which questions the previously predicted inhibitory activity against aspartic proteinases. Moreover, Egyptian cotton worm (Spodoptera littoralis) larvae fed on transgenic tomato plants overexpressing JIP21 present an increase in mortality and a delay in growth when compared with larvae fed on wild-type plants. These larvae belong to the Lepidoptera family whose main digestive enzymes have been described as being Ser proteases. All these results support the notion that tomato JIP21 should be considered as a chymotrypsin inhibitor belonging to the Ser proteinase inhibitors rather than a CDI. Therefore, we propose to name this protein tomato chymotrypsin inhibitor 21 (TCI21).

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“…These include potato (Atkinson, 1996), rice (Vain et al, 1998), Cavendish banana (Atkinson et al, 2004) for nematodes and potato (Lecardonnel et al, 1999) for coleopterans. In all these cases, increased protection to nematode and coleopteran damage was observed.…”

Section: Resultsmentioning

confidence: 99%

Agrobacterium mediated transformation of banana (Musa sp.) cv. Sukali Ndiizi (ABB) with a modified Carica papaya cystatin (CpCYS) gene

Namuddu¹,

Kiggundu²,

A³

et al. 2013

Afr. J. Biotechnol.

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Conventional banana breeding for pest and disease resistance is a very difficult and slow process due to the limited sources of resistance, sterility of cultivated banana varieties, high polyploidy levels, long cropping cycle and the lack of rapid screening methods. Molecular breeding using the transgenic approach with candidate genes such as cystatins offers an alternative method to banana improvement. Cystatin proteins inhibit the activity of cysteine proteases responsible for the breakdown of dietary proteins in the gut of many pests including nematodes resulting in protein deficiency. In this study, the papaya cystatin gene was introduced into the banana genome. Embryogenic cell suspension (ECS) cultures of the banana cultivar Sukali Ndiizi (ABB) were used as explants material for the successful transformation of banana. The Carica papaya cystatin gene (CpCYS-Mut89) previously modified to improve its inhibitory potential against banana pests was introduced into this cultivar using Agrobacterium tumefaciens, strain LBA4404 and the gus reporter gene was used to observe successful transformation process. We report the successful protocol for routine transformation of this cultivar, which was completed in six months with plant regeneration observed at a frequency of 23%. An additional four months was required to multiply the regenerant lines in order to have at least 20 plants per line for downstream challenging studies. Putatively transgenic plants were analyzed by PCR using hpt and CpCYS-Mut89 specific primers to confirm the presence of transgenes. Out of 28 selected lines, 27 were positive for both hpt and CpCYS-Mut89 transgenes giving 96.4% transformation efficiency. Five lines were then selected on the basis of putative PCR positives and a Southern blot analysis gave hybridization signals with 1 to 4 copy number integration patterns characteristic of Agrobacterium mediated transformation. These results confirm stable gene integration in East African banana cultivar cv. Sukali Ndiizi (genome group ABB) through an efficient Agrobacterium-mediated transformation protocol described for routine use in future improvement of this crop with genes of economic importance.

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Effects of rice cystatin I expression in transgenic potato on Colorado potato beetle larvae

Cited by 91 publications

References 39 publications

Tailoring the Specificity of a Plant Cystatin toward Herbivorous Insect Digestive Cysteine Proteases by Single Mutations at Positively Selected Amino Acid Sites

Tailoring the Specificity of a Plant Cystatin toward Herbivorous Insect Digestive Cysteine Proteases by Single Mutations at Positively Selected Amino Acid Sites

A Novel Function for the Cathepsin D Inhibitor in Tomato

Agrobacterium mediated transformation of banana (Musa sp.) cv. Sukali Ndiizi (ABB) with a modified Carica papaya cystatin (CpCYS) gene

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