Cyclotide Interactions with the Nematode External Surface

Abstract

Cyclotides are a large family of cyclic cystine knot-containing plant peptides that have anthelminthic activities against Haemonchus contortus and Trichostrongylus colubriformis, two important gastrointestinal nematodes of sheep. In this study, we investigated the interaction of the prototypic cyclotide kalata B1 with the external surface of H. contortus larvae and adult worms. We show that cyclotides do not need to be ingested by the worms to exert their toxic effects but that an interaction with the external surface alone is toxic. Evidence for this was the toxicity toward adult worms in the presence of a chemically induced pharyngeal ligature and toxicity of cyclotides toward nonfeeding larval life stages. Uptake of tritiated inulin in ligated adult worms was increased in the presence of cyclotide, suggesting that cyclotides increase the permeability of the external membranes of adult nematodes. Polyethylene glycols of various sizes showed protective effects on the nonfeeding larval life stage, as well as in hemolytic activity assays, suggesting that discrete pores are formed in the membrane surfaces by cyclotides and that these can be blocked by polyethylene glycols of appropriate size. This increased permeability is consistent with recently reported effects of cyclotides on membranes in which kalata B1 was demonstrated to form pores and cause leakage of vesicle/cellular contents. Our data, together with known size constraints on the movement of permeants across nematode cuticle layers, suggest that one action of the cyclotides involves an interaction with the lipid-rich epicuticle layer at the surface of the worm.Cyclotides are disulfide-rich peptides from plants that are characterized by their cyclic peptide backbone; they are the largest known family of genetically encoded cyclic proteins (11,23,46). More than 140 cyclotides have been reported to exist in the Violaceae and Rubiaceae plant families, and this number is expanding rapidly. Recent investigations suggest that thousands more cyclotides have yet to be discovered (46). Cyclotides exhibit a diverse range of biological activities, including anti-HIV (17, 22), antimicrobial (50), antineurotensin (52), cytotoxic (28, 49), and hemolytic (3, 40) activities. It has been postulated that the natural function of cyclotides is one of defense against plant pests and pathogens (2,23,24). The cyclotides kalata B1 (kB1) and kalata B2 (kB2) have a dramatic effect on growth and development of Helicoverpa punctigera (23) and Helicoverpa armigera larvae (24), two major pests of cotton. Recently, we demonstrated that cyclotides isolated from the plant Oldenlandia affinis display anthelminthic activity toward the gastrointestinal parasitic nematodes Haemonchus contortus and Trichostrongylus colubriformis (8). Subsequent studies found that cycloviolacins (a group of cyclotides isolated from Viola odorata) showed up to an 18-foldincreased potency against these nematodes relative to the prototypic cyclotide, kB1 (9). More recently, selected cyclotides have been shown to ...

Cyclotides, a Novel Ultrastable Polypeptide Scaffold for Drug Discovery

“…The natural function of cyclotides in plants seems to be as host-defense agents as deduced from their activity against insects [21, 22], nematodes [28, 29, 113, 114], and mollusks [60]. For example, it has been shown that cyclotides can efficiently inhibit the growth and development of insect and nematode larvae [115].…”

The bountiful biological activities of cyclotides

“…3 | Issue 3 | Jul-Sep 2012  172  novel drugs and biopesticides. [34,35] The speculated natural function of cyclotides is in plant defense as illustrated by several reports of their antibacterial/antimicrobial, [36] insecticidal, [37,38] antihelmintic, [39][40][41] nematocidal, [42] antifouling, [43] and molluscicidal properties. [44] The use of cyclotides in human health applications was first explored during the discovery of kalata B1, and although its uterotonic activity was established in rat, rabbit, and human uteri, it is not recommended as an oxytocic agent because of the severe side effects.…”

“…[47,50,52,[69][70][71][72][73] For instance, reports indicate that kalata peptides selectively bind to bacterial membranes, [69,74] and some cyclotides may form discrete pores on the external surface of nematodes and thereby interact with the lipid-rich epicuticle layer at the surface of the worm. [42] Furthermore, the antitumor properties of cyclotide, cycloviolacin O 2 (CyO 2 ) are caused first by the disruption of lipid membranes followed by leakage of contents from whole cells as well as liposomes. [70] CyO 2 causes rapid (within 5 minutes) disruption of lipid bilayers and is selectively cytotoxic in a dose-dependent manner.…”

Cyclotides

“…56 These results indicated the structural regions involved in nematicidal activity and suggested that the inclusion of epitopes in loops helps in interaction with the membrane. 56,57 Antimicrobial Activity Bactericidal activity is an important function related to the cyclotide family. The cyclotides circulin B and cyclopsychotride A show antimicrobial activities against Escherichia coli and Pseudomonas aeruginosa.…”