Biological Effects of Canatoxin in Different Insect Models: Evidence for a Proteolytic Activation of the Toxin by Insect Cathepsinlike Enzymes

Abstract

Canatoxin is a toxic protein isolated from the jackbean, Canavalia ensiformis. The toxin injected intraperitoneally is lethal for mice and rats; however, it is inactive if given orally. In this study, Manduca sexta (L.) (Lepidoptera), Schistocerca americana (Drury) (Orthoptera), Drosophila melanogaster (L.) (Diptera), Aedes aegypti (L.) (Diptera), Rhodnius prolixus (Stål) (Hemiptera), and Callosobruchus maculatus (F.) (Coleoptera) were fed on canatoxin-containing diets. No effects were seen in M. sexta, S. americana, D. melanogaster, or A. aegypti. No traces of canatoxin were found in their feces, suggesting that the protein was digested completely by these insects, which characteristically have a trypsin-based digestion. In contrast, canatoxin was lethal for insects displaying cathepsin-based digestion. Thus, for C. maculatus, a diet containing 0.25% wt:wt canatoxin caused complete inhibition of larval growth. When R. prolixus were fed on canatoxin, 2 effects were seen: impairment of water excretion and increased lethality 48-96 h after feeding. The lethal effect of canatoxin in R. prolixus was blocked partially or completely when the digestion of the toxin by R. prolixus midgut enzymes was impaired. The data showed that canatoxin is highly toxic when ingested by some species of insects but not affecting others, probably in correlation with the characteristics of the digestive process of the insect.

Stage-specific gut proteinases of the cotton stainer bug Dysdercus peruvianus: Role in the release of entomotoxic peptides from Canavalia ensiformis urease

“…In plants, ureases are homotrimers or hexamers of a w90 kDa subunit and participate in the use of urea as nitrogen source (Polacco and Holland, 1993;Sirko and Brodzik, 2000;Follmer, 2008). Canatoxin, a toxic protein isolated from Canavalia ensiformis seeds (Carlini and Guimarães, 1981) and more recently identified as a minor isoform of urease (Follmer et al, 2001), displays insecticidal properties (Carlini et al, 1997). Canatoxin and the major isoform of urease of C. ensiformis seeds (herein designated ''urease'') are toxic to Dysdercus peruvianus (Hemiptera: Pyrrhocoridae), and this toxicity is independent of their ureolytic activity (Follmer et al, 2004a;Stanisçuaski et al, 2005;Follmer et al, 2004b).…”

Silencing urease: A key evolutionary step that facilitated the adaptation of Yersinia pestis to the flea-borne transmission route

“…Mice and rats have high blood urea nitrogen levels (2-3 mg/mL). Interestingly, jack bean and soybean ureases have been shown previously to have toxic effects when fed to insects (26,27). However, insect toxicity of these plant ureases has nothing to do with enzymatic activity, but is due to a 10-kDa toxic peptide fragment released from the urease protein by insect cathepsinlike proteases.…”

Ureases display biological effects independent of enzymatic activity: Is there a connection to diseases caused by urease-producing bacteria?

“…The insecticidal properties of plant ureases were first described for canatoxin (12) and later for C. ensiformis major urease and soybean embryo-specific urease (13). The kissing bug Rhodnius prolixus, and three economically important crop pests, the cowpea weevil Callosobruchus maculatus, the green stinkbug Nezara viridula and the cotton stainer bug Dysdercus peruvianus, are susceptible to the lethal effect of these proteins when they are added to their diets at 0.02 to 0.1% (w/w) levels (14,15).…”

“…Canatoxin and urease are hydrolyzed by these enzymes to release an internal entomotoxic peptide of 10 kDa (16). No effects of intact canatoxin/ urease were seen in insects relying on trypsin-like digestive enzymes, which apparently degrade the proteins more extensively (12). A recombinant peptide, equivalent to that produced by hydrolysis of canatoxin with insect cathepsins, was obtained by heterologous expression in Escherichia coli.…”