Effects of potato plants expressing a barley cystatin on the predatory bug Podisus maculiventris via herbivorous prey feeding on the plant

Abstract

The aim of this study was to assess the effects of potato plants expressing a barley cystatin on a potentially cystatin-susceptible natural enemy by predation on susceptible and non-susceptible preys feeding on the plant. We have focussed on the impact of the variant HvCPI-1 C68 --> G, in which the only cysteine residue was changed by a glycine, on the growth and digestive physiology of the Colorado potato beetle (CPB), Leptinotarsa decemlineata, and the Egyptian cotton leafworm (ECW), Spodoptera littoralis. Moreover, we have studied the prey-mediated effects of the barley cystatin at the third trophic level, using the predatory spined soldier bug (SSB), Podisus maculiventris, as a model. Feeding trials conducted with CPB larvae reared on transgenic potato plants expressing the C68 --> G variant resulted in significantly lower weight gains compared to those fed on non-transformed (NT) plants. On the contrary, larger weight gains were obtained when ECW larvae, that lack digestive cysteine proteases, were reared on transgenic potato expressing the cystatin, as compared to larvae fed on NT plants. No negative effects on survival and growth were observed when SSB nymphs were exposed to HvCPI-1 C68 --> G by predation on either CPB or ECW larvae reared on transgenic potato plants expressing the barley cystatin, despite the fact that the inhibitor suppressed in vitro gut proteolysis of the predatory bug. To investigate the physiological background, biochemical analysis were carried out on guts of insects dissected at the end of the feeding assays.

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

“…However, cathepsin D-and leucine aminopeptidase-like activities were not affected, suggesting that the spider mite did not develop a compensatory response to the inhibitory effect mediated by the barley cystatin. The overproduction of non-target proteases is usually found in other phytophagous arthropods as a response to host plants expressing distinct sets of defence proteins (Lara et al 2000;Alvarez-Alfageme et al 2007). …”

“…Leaves were ground and resuspended in 0.15 M NaCl sodium phosphate pH 6.0, 2 mM EDTA for 1 h at 4°C and treated as described in Alvarez-Alfageme et al (2007). Inhibitory activity of plant protein extracts (20 ug) was in vitro tested against 10 ng of commercial papain (EC 3.4.22.2) from Sigma using Z-FR-AMC as substrate.…”

“…From a functional viewpoint, PhyCys have been implicated in regulation of the protein turn-over and as defense proteins. The defense role has been inferred from the ability of PhyCys to inhibit digestive proteases from herbivorous arthropods in vitro, as well as by bioassays in artificial diets and on transgenic plants over-expressing PhyCys genes (Pernas et al 1998;Alvarez-Alfageme et al 2007;Carrillo et al 2010). Deleterious effects against phytopathogenic fungi and virases have also been described for these inhibitors (Gutiérrez-Campos et al 1999;Martinez et al 2003;Abraham et al 2006).…”

Deleterious effects of plant cystatins against the banana weevil Cosmopolites sordidus

“…Despite some promising developments (Leplé et al, 1995;Kuroda et al, 1996;Koiwa et al, 2000;Liu et al, 2004;Alvarez-Alfageme et al, 2007;Ninkovic et al, 2007), the general usefulness of plant cystatins for the control of coleopteran pests still remains to be established. These insects have developed over time effective strategies to elude the inhibitory effects of plant protease inhibitors, involving the use of complex digestive protease systems with proteases from different mechanistic classes acting in a complementary, coordinated manner (Brunelle et al, 1999;Hernandez et al, 2003;Gruden et al, 2003;Vinokurov et al, 2006a,b;Prabhakar et al, 2007); the over-expression of target proteases following cystatin ingestion to outnumber the inhibitory proteins (Cloutier et al, 2000;Ahn et al, 2004); the constitutive or diet-induced expression of cysteine cathepsins weakly sensitive to the ingested cystatin, the so-called 'cystatin-insensitive proteases' (Michaud et al, 1993(Michaud et al, , 1995aGirard et al, 1998a;Cloutier et al, 1999Cloutier et al, , 2000ZhuSalzman et al, 2003;Brunelle et al, 2004;Gruden et al, 2004;Liu et al, 2004;Koo et al, 2008); the over-expression of proteases from alternative mechanistic classes following cystatin ingestion (Zhu-Salzman et al, 2003;Brunelle et al, 2004;Rivard et al, 2004;Oppert et al, 2005); and the degradation of defensive protease inhibitors using non-target, insensitive proteases (Michaud et al, 1995b;Girard et al, 1998b;Giri et al, 1998;Gruden et al, 2003;Zhu-Salzman et al, 2003).…”

“…In this study, we assessed the potential of OC-I and other plant cystatins as possible tools for the production of transgenic banana lines resistant to the banana weevil. Several papers have been published assessing the effectiveness of plant cystatins against herbivorous Coleoptera, with conclusive results in some cases (Leplé et al, 1995;Kuroda et al, 1996;Lecardonnel et al, 1999;Koiwa et al, 2000;Alvarez-Alfageme et al 2007;Ninkovic et al, 2007) but disappointing results in other cases owing to the ability of the target insects to readily elude or even counteract the antidigestive effects of natural or recombinant inhibitors (e.g. Girard et al, 1998a,b;Cloutier et al, 1999Cloutier et al, , 2000Zhu-Salzman et al, 2003;Gruden et al, 2003Gruden et al, , 2004Ahn et al, 2004Ahn et al, , 2007Oppert et al, 2005;Koo et al, 2008).…”