Prey Shifts Drive Venom Evolution in Cone Snails.

Venom systems are complex traits that have independently emerged multiple times in diverse plant and animal phyla. Within each venomous lineage there typically exists interspecific variation in venom composition where several factors have been proposed as drivers of variation, including phylogeny and diet. Understanding these factors is of broad biological interest and has implications for the development of antivenom therapies and venom-based drug discovery. Because of their high species richness and the presence of several major evolutionary prey shifts, venomous marine cone snails (genus Conus) provide an ideal system to investigate drivers of interspecific venom variation. Here, by analyzing the venom gland expression profiles of ∼3,000 toxin genes from 42 species of cone snail, we elucidate the role of prey-specific selection pressures in shaping venom variation. By analyzing overall venom composition and individual toxin structures, we demonstrate that the shifts from vermivory to piscivory in Conus are complemented by distinct changes in venom composition independent of phylogeny. In vivo injections of venom from piscivorous cone snails in fish further showed a higher potency compared with venom of nonpiscivores demonstrating a selective advantage. Together, our findings provide compelling evidence for the role of prey shifts in directing the venom composition of cone snails and expand our understanding of the mechanisms of venom variation and diversification.

Analogs of α-conotoxin PnIC selectively inhibit α7ß2- over α7-only subtype nicotinic acetylcholine receptors via a novel allosteric mechanism.

This study was undertaken to identify and characterize the first ligands capable of selectively identifying nicotinic acetylcholine receptors containing α7 and ß2 subunits (α7ß2-nAChR subtype). Basal forebrain cholinergic neurons express α7ß2-nAChR. Here, they appear to mediate neuronal dysfunction induced by the elevated levels of oligomeric amyloid-ß associated with early Alzheimer's disease. Additional work indicates that α7ß2-nAChR are expressed across several further critically important cholinergic and GABAergic neuronal circuits within the central nervous system. Further studies, however, are significantly hindered by the inability of currently available ligands to distinguish heteromeric α7ß2-nAChR from the closely related and more widespread homomeric α7-only-nAChR subtype. Functional screening using two-electrode voltage-clamp electrophysiology identified a family of α7ß2-nAChR-selective analogs of α-conotoxin PnIC (α-CtxPnIC). A combined electrophysiology, functional kinetics, site-directed mutagenesis, and molecular dynamics approach was used to further characterize the α7ß2-nAChR selectivity and site of action of these α-CtxPnIC analogs. We determined that α7ß2-nAChR selectivity of α-CtxPnIC analogs arises from interactions at a site distinct from the orthosteric agonist-binding site shared between α7ß2- and α7-only-nAChR. As numerous previously identified α-Ctx ligands are competitive antagonists of orthosteric agonist-binding sites, this study profoundly expands the scope of use of α-Ctx ligands (which have already provided important nAChR research and translational breakthroughs). More immediately, analogs of α-CtxPnIC promise to enable, for the first time, both comprehensive mapping of the distribution of α7ß2-nAChR and detailed investigations of their physiological roles.

Using Constellation Pharmacology to Characterize a Novel α-Conotoxin from Conus ateralbus.

The venom of cone snails has been proven to be a rich source of bioactive peptides that target a variety of ion channels and receptors. α-Conotoxins (αCtx) interact with nicotinic acetylcholine receptors (nAChRs) and are powerful tools for investigating the structure and function of the various nAChR subtypes. By studying how conotoxins interact with nAChRs, we can improve our understanding of these receptors, leading to new insights into neurological diseases associated with nAChRs. Here, we describe the discovery and characterization of a novel conotoxin from Conus ateralbus, αCtx-AtIA, which has an amino acid sequence homologous to the well-described αCtx-PeIA, but with a different selectivity profile towards nAChRs. We tested the synthetic αCtx-AtIA using the calcium imaging-based Constellation Pharmacology assay on mouse DRG neurons and found that αCtx-AtIA significantly inhibited ACh-induced calcium influx in the presence of an α7 positive allosteric modulator, PNU-120596 (PNU). However, αCtx-AtIA did not display any activity in the absence of PNU. These findings were further validated using two-electrode voltage clamp electrophysiology performed on oocytes overexpressing mouse α3ß4, α6/α3ß4 and α7 nAChRs subtypes. We observed that αCtx-AtIA displayed no or low potency in blocking α3ß4 and α6/α3ß4 receptors, respectively, but improved potency and selectivity to block α7 nAChRs when compared with αCtx-PeIA. Through the synthesis of two additional analogs of αCtx-AtIA and subsequent characterization using Constellation Pharmacology, we were able to identify residue Trp18 as a major contributor to the activity of the peptide.

Fish-hunting cone snail disrupts prey's glucose homeostasis with weaponized mimetics of somatostatin and insulin.

Venomous animals have evolved diverse molecular mechanisms to incapacitate prey and defend against predators. Most venom components disrupt nervous, locomotor, and cardiovascular systems or cause tissue damage. The discovery that certain fish-hunting cone snails use weaponized insulins to induce hypoglycemic shock in prey highlights a unique example of toxins targeting glucose homeostasis. Here, we show that, in addition to insulins, the deadly fish hunter, Conus geographus, uses a selective somatostatin receptor 2 (SSTR2) agonist that blocks the release of the insulin-counteracting hormone glucagon, thereby exacerbating insulin-induced hypoglycemia in prey. The native toxin, Consomatin nG1, exists in several proteoforms with a minimized vertebrate somatostatin-like core motif connected to a heavily glycosylated N-terminal region. We demonstrate that the toxin's N-terminal tail closely mimics a glycosylated somatostatin from fish pancreas and is crucial for activating the fish SSTR2. Collectively, these findings provide a stunning example of chemical mimicry, highlight the combinatorial nature of venom components, and establish glucose homeostasis as an effective target for prey capture.