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1.
PLoS Biol ; 21(8): e3002217, 2023 08.
Artículo en Inglés | MEDLINE | ID: mdl-37535677

RESUMEN

Animal venom peptides represent valuable compounds for biomedical exploration. The venoms of marine cone snails constitute a particularly rich source of peptide toxins, known as conotoxins. Here, we identify the sequence of an unusually large conotoxin, Mu8.1, which defines a new class of conotoxins evolutionarily related to the well-known con-ikot-ikots and 2 additional conotoxin classes not previously described. The crystal structure of recombinant Mu8.1 displays a saposin-like fold and shows structural similarity with con-ikot-ikot. Functional studies demonstrate that Mu8.1 curtails calcium influx in defined classes of murine somatosensory dorsal root ganglion (DRG) neurons. When tested on a variety of recombinantly expressed voltage-gated ion channels, Mu8.1 displayed the highest potency against the R-type (Cav2.3) calcium channel. Ca2+ signals from Mu8.1-sensitive DRG neurons were also inhibited by SNX-482, a known spider peptide modulator of Cav2.3 and voltage-gated K+ (Kv4) channels. Our findings highlight the potential of Mu8.1 as a molecular tool to identify and study neuronal subclasses expressing Cav2.3. Importantly, this multidisciplinary study showcases the potential of uncovering novel structures and bioactivities within the largely unexplored group of macro-conotoxins.


Asunto(s)
Conotoxinas , Ratones , Animales , Conotoxinas/farmacología , Conotoxinas/química , Canales de Calcio , Péptidos/química , Células Receptoras Sensoriales/metabolismo , Caracoles
2.
Mol Biol Evol ; 41(8)2024 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-38935574

RESUMEN

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.


Asunto(s)
Caracol Conus , Venenos de Moluscos , Animales , Caracol Conus/genética , Venenos de Moluscos/genética , Conducta Predatoria , Evolución Biológica , Filogenia , Evolución Molecular
3.
Nat Chem Biol ; 18(7): 688-697, 2022 07.
Artículo en Inglés | MEDLINE | ID: mdl-35761080

RESUMEN

Insulin and its related peptides are found throughout the animal kingdom, in which they serve diverse functions. This includes regulation of glucose homeostasis, neuronal development and cognition. The surprising recent discovery that venomous snails evolved specialized insulins to capture fish demonstrated the nefarious use of this hormone in nature. Because of their streamlined role in predation, these repurposed insulins exhibit unique characteristics that have unraveled new aspects of the chemical ecology and structural biology of this important hormone. Recently, insulins were also reported in other venomous predators and pathogenic viruses, demonstrating the broader use of insulin by one organism to manipulate the physiology of another. In this Review, we provide an overview of the discovery and biomedical application of repurposed insulins and other hormones found in nature and highlight several unique insights gained from these unusual compounds.


Asunto(s)
Insulina , Insulinas , Animales
4.
Nat Chem Biol ; 18(5): 511-519, 2022 05.
Artículo en Inglés | MEDLINE | ID: mdl-35289328

RESUMEN

Cone snail venoms contain a wide variety of bioactive peptides, including insulin-like molecules with distinct structural features, binding modes and biochemical properties. Here, we report an active humanized cone snail venom insulin with an elongated A chain and a truncated B chain, and use cryo-electron microscopy (cryo-EM) and protein engineering to elucidate its interactions with the human insulin receptor (IR) ectodomain. We reveal how an extended A chain can compensate for deletion of B-chain residues, which are essential for activity of human insulin but also compromise therapeutic utility by delaying dissolution from the site of subcutaneous injection. This finding suggests approaches to developing improved therapeutic insulins. Curiously, the receptor displays a continuum of conformations from the symmetric state to a highly asymmetric low-abundance structure that displays coordination of a single humanized venom insulin using elements from both of the previously characterized site 1 and site 2 interactions.


Asunto(s)
Insulina , Venenos de Moluscos , Microscopía por Crioelectrón , Humanos , Insulina/metabolismo , Venenos de Moluscos/química , Venenos de Moluscos/metabolismo , Péptidos , Conformación Proteica
5.
Cell Mol Life Sci ; 80(10): 287, 2023 Sep 09.
Artículo en Inglés | MEDLINE | ID: mdl-37689602

RESUMEN

Voltage-gated sodium (NaV) channels are transmembrane proteins that play a critical role in electrical signaling in the nervous system and other excitable tissues. µ-Conotoxins are peptide toxins from the venoms of marine cone snails (genus Conus) that block NaV channels with nanomolar potency. Most species of the subgenera Textilia and Afonsoconus are difficult to acquire; therefore, their venoms have yet to be comprehensively interrogated for µ-conotoxins. The goal of this study was to find new µ-conotoxins from species of the subgenera Textilia and Afonsoconus and investigate their selectivity at human NaV channels. Using RNA-seq of the venom gland of Conus (Textilia) bullatus, we identified 12 µ-conotoxin (or µ-conotoxin-like) sequences. Based on these sequences we designed primers which we used to identify additional µ-conotoxin sequences from DNA extracted from historical specimens of species from Textilia and Afonsoconus. We synthesized six of these µ-conotoxins and tested their activity on human NaV1.1-NaV1.8. Five of the six synthetic peptides were potent blockers of human NaV channels. Of these, two peptides (BuIIIB and BuIIIE) were potent blockers of hNaV1.3. Three of the peptides (BuIIIB, BuIIIE and AdIIIA) had submicromolar activity at hNaV1.7. This study serves as an example of the identification of new peptide toxins from historical DNA and provides new insights into structure-activity relationships of µ-conotoxins with activity at hNaV1.3 and hNaV1.7.


Asunto(s)
Conotoxinas , Caracol Conus , Toxinas Biológicas , Humanos , Animales , Conotoxinas/farmacología , Proteínas de la Membrana , Canales de Sodio/genética
6.
Mol Biol Evol ; 39(4)2022 04 10.
Artículo en Inglés | MEDLINE | ID: mdl-35383850

RESUMEN

Somatostatin and its related peptides (SSRPs) form an important family of hormones with diverse physiological roles. The ubiquitous presence of SSRPs in vertebrates and several invertebrate deuterostomes suggests an ancient origin of the SSRP signaling system. However, the existence of SSRP genes outside of deuterostomes has not been established, and the evolutionary history of this signaling system remains poorly understood. Our recent discovery of SSRP-like toxins (consomatins) in venomous marine cone snails (Conus) suggested the presence of a related signaling system in mollusks and potentially other protostomes. Here, we identify the molluscan SSRP-like signaling gene that gave rise to the consomatin family. Following recruitment into venom, consomatin genes experienced strong positive selection and repeated gene duplications resulting in the formation of a hyperdiverse family of venom peptides. Intriguingly, the largest number of consomatins was found in worm-hunting species (>400 sequences), indicating a homologous system in annelids, another large protostome phylum. Consistent with this, comprehensive sequence mining enabled the identification of SSRP-like sequences (and their corresponding orphan receptor) in annelids and several other protostome phyla. These results established the existence of SSRP-like peptides in many major branches of bilaterians and challenge the prevailing hypothesis that deuterostome SSRPs and protostome allatostatin-C are orthologous peptide families. Finally, having a large set of predator-prey SSRP sequences available, we show that although the cone snail's signaling SSRP-like genes are under purifying selection, the venom consomatin genes experience rapid directional selection to target receptors in a changing mix of prey.


Asunto(s)
Conotoxinas , Caracol Conus , Animales , Conotoxinas/genética , Caracol Conus/genética , Neuropéptidos , Péptidos/genética , Somatostatina/genética , Ponzoñas
7.
Mar Drugs ; 21(2)2023 Jan 25.
Artículo en Inglés | MEDLINE | ID: mdl-36827123

RESUMEN

Elevenins are peptides found in a range of organisms, including arthropods, annelids, nematodes, and molluscs. They consist of 17 to 19 amino acid residues with a single conserved disulfide bond. The subject of this study, elevenin-Vc1, was first identified in the venom of the cone snail Conus victoriae (Gen. Comp. Endocrinol. 2017, 244, 11-18). Although numerous elevenin sequences have been reported, their physiological function is unclear, and no structural information is available. Upon intracranial injection in mice, elevenin-Vc1 induced hyperactivity at doses of 5 or 10 nmol. The structure of elevenin-Vc1, determined using nuclear magnetic resonance spectroscopy, consists of a short helix and a bend region stabilised by the single disulfide bond. The elevenin-Vc1 structural fold is similar to that of α-conotoxins such as α-RgIA and α-ImI, which are also found in the venoms of cone snails and are antagonists at specific subtypes of nicotinic acetylcholine receptors (nAChRs). In an attempt to mimic the functional motif, Asp-Pro-Arg, of α-RgIA and α-ImI, we synthesised an analogue, designated elevenin-Vc1-DPR. However, neither elevenin-Vc1 nor the analogue was active at six different human nAChR subtypes (α1ß1εδ, α3ß2, α3ß4, α4ß2, α7, and α9α10) at 1 µM concentrations.


Asunto(s)
Conotoxinas , Caracol Conus , Receptores Nicotínicos , Ratones , Humanos , Animales , Conotoxinas/farmacología , Caracol Conus/metabolismo , Ponzoñas , Receptores Nicotínicos/metabolismo , Péptidos/metabolismo , Antagonistas Nicotínicos/farmacología
8.
J Nat Prod ; 84(4): 1232-1243, 2021 04 23.
Artículo en Inglés | MEDLINE | ID: mdl-33764053

RESUMEN

Natural products such as conotoxins have tremendous potential as tools for biomedical research and for the treatment of different human diseases. Conotoxins are peptides present in the venoms of predatory cone snails that have a rich diversity of pharmacological functions. One of the major bottlenecks in natural products research is the rapid identification and evaluation of bioactive molecules. To overcome this limitation, we designed a set of light-induced behavioral assays in zebrafish larvae to screen for bioactive conotoxins. We used this screening approach to test several unique conotoxins derived from different cone snail clades and discovered that a conorfamide from Conus episcopatus, CNF-Ep1, had the most dramatic alterations in the locomotor behavior of zebrafish larvae. Interestingly, CNF-Ep1 is also bioactive in several mouse assay systems when tested in vitro and in vivo. Our novel screening platform can thus accelerate the identification of bioactive marine natural products, and the first compound discovered using this assay has intriguing properties that may uncover novel neuronal circuitry.


Asunto(s)
Larva/efectos de los fármacos , Locomoción/efectos de los fármacos , Venenos de Moluscos/farmacología , Neuropéptidos/farmacología , Pez Cebra , Animales , Caracol Conus/química , Femenino , Masculino , Ratones
9.
J Biol Chem ; 294(22): 8745-8759, 2019 05 31.
Artículo en Inglés | MEDLINE | ID: mdl-30975904

RESUMEN

Venomous marine cone snails produce peptide toxins (conotoxins) that bind ion channels and receptors with high specificity and therefore are important pharmacological tools. Conotoxins contain conserved cysteine residues that form disulfide bonds that stabilize their structures. To gain structural insight into the large, yet poorly characterized conotoxin H-superfamily, we used NMR and CD spectroscopy along with MS-based analyses to investigate H-Vc7.2 from Conus victoriae, a peptide with a VI/VII cysteine framework. This framework has CysI-CysIV/CysII-CysV/CysIII-CysVI connectivities, which have invariably been associated with the inhibitor cystine knot (ICK) fold. However, the solution structure of recombinantly expressed and purified H-Vc7.2 revealed that although it displays the expected cysteine connectivities, H-Vc7.2 adopts a different fold consisting of two stacked ß-hairpins with opposing ß-strands connected by two parallel disulfide bonds, a structure homologous to the N-terminal region of the human granulin protein. Using structural comparisons, we subsequently identified several toxins and nontoxin proteins with this "mini-granulin" fold. These findings raise fundamental questions concerning sequence-structure relationships within peptides and proteins and the key determinants that specify a given fold.


Asunto(s)
Conotoxinas/química , Caracol Conus/metabolismo , Cisteína/química , Granulinas/química , Secuencia de Aminoácidos , Animales , Conotoxinas/genética , Conotoxinas/metabolismo , Disulfuros/química , Granulinas/metabolismo , Espectroscopía de Resonancia Magnética , Venenos de Moluscos/metabolismo , Conformación Proteica en Lámina beta , Pliegue de Proteína , Estabilidad Proteica , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/química , Proteínas Recombinantes/genética
10.
Proc Natl Acad Sci U S A ; 113(12): 3227-32, 2016 Mar 22.
Artículo en Inglés | MEDLINE | ID: mdl-26957604

RESUMEN

Formation of correct disulfide bonds in the endoplasmic reticulum is a crucial step for folding proteins destined for secretion. Protein disulfide isomerases (PDIs) play a central role in this process. We report a previously unidentified, hypervariable family of PDIs that represents the most diverse gene family of oxidoreductases described in a single genus to date. These enzymes are highly expressed specifically in the venom glands of predatory cone snails, animals that synthesize a remarkably diverse set of cysteine-rich peptide toxins (conotoxins). Enzymes in this PDI family, termed conotoxin-specific PDIs, significantly and differentially accelerate the kinetics of disulfide-bond formation of several conotoxins. Our results are consistent with a unique biological scenario associated with protein folding: The diversification of a family of foldases can be correlated with the rapid evolution of an unprecedented diversity of disulfide-rich structural domains expressed by venomous marine snails in the superfamily Conoidea.


Asunto(s)
Venenos de Moluscos/química , Péptidos/química , Proteína Disulfuro Isomerasas/genética , Secuencia de Aminoácidos , Animales , Caracol Conus , Datos de Secuencia Molecular , Proteína Disulfuro Isomerasas/química , Pliegue de Proteína , Homología de Secuencia de Aminoácido
11.
Proc Natl Acad Sci U S A ; 112(6): 1743-8, 2015 Feb 10.
Artículo en Inglés | MEDLINE | ID: mdl-25605914

RESUMEN

More than 100 species of venomous cone snails (genus Conus) are highly effective predators of fish. The vast majority of venom components identified and functionally characterized to date are neurotoxins specifically targeted to receptors, ion channels, and transporters in the nervous system of prey, predators, or competitors. Here we describe a venom component targeting energy metabolism, a radically different mechanism. Two fish-hunting cone snails, Conus geographus and Conus tulipa, have evolved specialized insulins that are expressed as major components of their venoms. These insulins are distinctive in having much greater similarity to fish insulins than to the molluscan hormone and are unique in that posttranslational modifications characteristic of conotoxins (hydroxyproline, γ-carboxyglutamate) are present. When injected into fish, the venom insulin elicits hypoglycemic shock, a condition characterized by dangerously low blood glucose. Our evidence suggests that insulin is specifically used as a weapon for prey capture by a subset of fish-hunting cone snails that use a net strategy to capture prey. Insulin appears to be a component of the nirvana cabal, a toxin combination in these venoms that is released into the water to disorient schools of small fish, making them easier to engulf with the snail's distended false mouth, which functions as a net. If an entire school of fish simultaneously experiences hypoglycemic shock, this should directly facilitate capture by the predatory snail.


Asunto(s)
Caracol Conus/química , Caracol Conus/fisiología , Insulina/genética , Toxinas Marinas/química , Conducta Predatoria/fisiología , Pez Cebra/metabolismo , Secuencia de Aminoácidos , Animales , Insulina/análisis , Insulina/síntesis química , Insulina/metabolismo , Toxinas Marinas/metabolismo , Espectrometría de Masas , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Alineación de Secuencia , Análisis de Secuencia de ADN , Especificidad de la Especie
12.
Proc Natl Acad Sci U S A ; 112(16): 5087-92, 2015 Apr 21.
Artículo en Inglés | MEDLINE | ID: mdl-25848010

RESUMEN

Prey shifts in carnivorous predators are events that can initiate the accelerated generation of new biodiversity. However, it is seldom possible to reconstruct how the change in prey preference occurred. Here we describe an evolutionary "smoking gun" that illuminates the transition from worm hunting to fish hunting among marine cone snails, resulting in the adaptive radiation of fish-hunting lineages comprising ∼100 piscivorous Conus species. This smoking gun is δ-conotoxin TsVIA, a peptide from the venom of Conus tessulatus that delays inactivation of vertebrate voltage-gated sodium channels. C. tessulatus is a species in a worm-hunting clade, which is phylogenetically closely related to the fish-hunting cone snail specialists. The discovery of a δ-conotoxin that potently acts on vertebrate sodium channels in the venom of a worm-hunting cone snail suggests that a closely related ancestral toxin enabled the transition from worm hunting to fish hunting, as δ-conotoxins are highly conserved among fish hunters and critical to their mechanism of prey capture; this peptide, δ-conotoxin TsVIA, has striking sequence similarity to these δ-conotoxins from piscivorous cone snail venoms. Calcium-imaging studies on dissociated dorsal root ganglion (DRG) neurons revealed the peptide's putative molecular target (voltage-gated sodium channels) and mechanism of action (inhibition of channel inactivation). The results were confirmed by electrophysiology. This work demonstrates how elucidating the specific interactions between toxins and receptors from phylogenetically well-defined lineages can uncover molecular mechanisms that underlie significant evolutionary transitions.


Asunto(s)
Caracol Conus/fisiología , Peces/fisiología , Conducta Predatoria/fisiología , Secuencia de Aminoácidos , Animales , Bioensayo , Conotoxinas/química , Conotoxinas/toxicidad , Caracol Conus/anatomía & histología , Datos de Secuencia Molecular , Péptidos/metabolismo , Filogenia
13.
Int J Mol Sci ; 19(11)2018 Oct 31.
Artículo en Inglés | MEDLINE | ID: mdl-30384459

RESUMEN

Disulfide-rich peptides are highly abundant in nature and their study has provided fascinating insight into protein folding, structure and function. Venomous cone snails belong to a group of organisms that express one of the largest sets of disulfide-rich peptides (conotoxins) found in nature. The diversity of structural scaffolds found for conotoxins suggests that specialized molecular adaptations have evolved to ensure their efficient folding and secretion. We recently showed that canonical protein disulfide isomerase (PDI) and a conotoxin-specific PDI (csPDI) are ubiquitously expressed in the venom gland of cone snails and play a major role in conotoxin folding. Here, we identify cone snail endoplasmic reticulum oxidoreductin-1 (Conus Ero1) and investigate its role in the oxidative folding of conotoxins through reoxidation of cone snail PDI and csPDI. We show that Conus Ero1 preferentially reoxidizes PDI over csPDI, suggesting that the reoxidation of csPDI may rely on an Ero1-independent molecular pathway. Despite the preferential reoxidation of PDI over csPDI, the combinatorial effect of Ero1 and csPDI provides higher folding yields than Ero1 and PDI. We further demonstrate that the highest in vitro folding rates of two model conotoxins are achieved when all three enzymes are present, indicating that these enzymes may act synergistically. Our findings provide new insight into the generation of one of the most diverse classes of disulfide-rich peptides and may improve current in vitro approaches for the production of venom peptides for pharmacological studies.


Asunto(s)
Conotoxinas/química , Caracol Conus/química , Oxidorreductasas/química , Proteína Disulfuro Isomerasas/química , Pliegue de Proteína , Animales , Oxidación-Reducción
14.
Biochim Biophys Acta ; 1864(6): 715-723, 2016 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-26947243

RESUMEN

Despite its critical role in maintaining glucose homeostasis, surprisingly little is known about proinsulin folding in the endoplasmic reticulum. In this study we aimed to understand the chaperones involved in the maturation and degradation of proinsulin. We generated pancreatic beta cell lines expressing FLAG-tagged proinsulin. Several chaperones (including BiP, PDIA6, calnexin, calreticulin, GRP170, Erdj3 and ribophorin II) co-immunoprecipitated with proinsulin suggesting a role for these proteins in folding. To investigate the chaperones responsible for targeting misfolded proinsulin for degradation, we also created a beta cell line expressing FLAG-tagged proinsulin carrying the Akita mutation (Cys96Tyr). All chaperones found to be associated with wild type proinsulin also co-immunoprecipitated with Akita proinsulin. However, one additional protein, namely P58(IPK), specifically precipitated with Akita proinsulin and approximately ten fold more PDIA6, but not other PDI family members, was bound to Akita proinsulin. The latter suggests that PDIA6 may act as a key reductase and target misfolded proinsulin to the ER-degradation pathway. The preferential association of PDIA6 to Akita proinsulin was also confirmed in another beta cell line (ßTC-6). Furthermore, for the first time, a physiologically relevant substrate for PDIA6 has been evidenced. Thus, this study has identified several chaperones/foldases that associated with wild type proinsulin and has also provided a comprehensive interactome for Akita misfolded proinsulin.


Asunto(s)
Proinsulina/química , Proteína Disulfuro Isomerasas/fisiología , Pliegue de Proteína , Animales , Línea Celular , Ratones , Mutagénesis Sitio-Dirigida , Proteína Disulfuro Isomerasas/química
15.
Mol Biol Evol ; 33(11): 2924-2934, 2016 11.
Artículo en Inglés | MEDLINE | ID: mdl-27524826

RESUMEN

A specialized insulin was recently found in the venom of a fish-hunting cone snail, Conus geographus Here we show that many worm-hunting and snail-hunting cones also express venom insulins, and that this novel gene family has diversified explosively. Cone snails express a highly conserved insulin in their nerve ring; presumably this conventional signaling insulin is finely tuned to the Conus insulin receptor, which also evolves very slowly. By contrast, the venom insulins diverge rapidly, apparently in response to biotic interactions with prey and also possibly the cones' own predators and competitors. Thus, the inwardly directed signaling insulins appear to experience predominantly purifying sele\ction to target an internal receptor that seldom changes, while the outwardly directed venom insulins frequently experience directional selection to target heterospecific insulin receptors in a changing mix of prey, predators and competitors. Prey insulin receptors may often be constrained in ways that prevent their evolutionary escape from targeted venom insulins, if amino-acid substitutions that result in escape also degrade the receptor's signaling functions.


Asunto(s)
Conotoxinas/genética , Caracol Conus/genética , Insulina/biosíntesis , Secuencia de Aminoácidos , Animales , Teorema de Bayes , Conotoxinas/biosíntesis , Conotoxinas/toxicidad , Caracol Conus/metabolismo , Evolución Molecular , Variación Genética , Insulina/genética , Datos de Secuencia Molecular , Receptor de Insulina/genética , Receptor de Insulina/metabolismo , Ponzoñas/biosíntesis , Ponzoñas/genética
16.
Artículo en Inglés | MEDLINE | ID: mdl-28551870

RESUMEN

From a biological perspective, a natural product can be defined as a compound evolved by an organism for chemical interactions with another organism including prey, predator, competitor, pathogen, symbiont or host. Natural products hold tremendous potential as drug leads and have been extensively studied by chemists and biochemists in the pharmaceutical industry. However, the biological purpose for which a natural product evolved is rarely addressed. By focusing on a well-studied group of natural products-venom components from predatory marine cone snails-this review provides a rationale for why a better understanding of the evolution, biology and biochemistry of natural products will facilitate both neuroscience and the potential for drug leads. The larger goal is to establish a new sub-discipline in the broader field of neuroethology that we refer to as "Chemical Neuroethology", linking the substantial work carried out by chemists on natural products with accelerating advances in neuroethology.


Asunto(s)
Evolución Biológica , Productos Biológicos/química , Caracol Conus/fisiología , Peces/fisiología , Conducta Predatoria/fisiología , Animales
17.
Gen Comp Endocrinol ; 244: 11-18, 2017 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-26301480

RESUMEN

The venoms of cone snails (genus Conus) are remarkably complex, consisting of hundreds of typically short, disulfide-rich peptides termed conotoxins. These peptides have diverse pharmacological targets, with injection of venom eliciting a range of physiological responses, including sedation, paralysis and sensory overload. Most conotoxins target the prey's nervous system but evidence of venom peptides targeting neuroendocrine processes is emerging. Examples include vasopressin, RFamide neuropeptides and recently also insulin. To investigate the diversity of hormone/neuropeptide-like molecules in the venoms of cone snails we systematically mined the venom gland transcriptomes of several cone snail species and examined secreted venom peptides in dissected and injected venom of the Australian cone snail Conus victoriae. Using this approach we identified several novel hormone/neuropeptide-like toxins, including peptides similar to the bee brain hormone prohormone-4, the mollusc ganglia neuropeptide elevenin, and thyrostimulin, a member of the glycoprotein hormone family, and confirmed the presence of insulin. We confirmed that at least two of these peptides are not only expressed in the venom gland but also form part of the injected venom cocktail, unambiguously demonstrating their role in envenomation. Our findings suggest that hormone/neuropeptide-like toxins are a diverse and integral part of the complex envenomation strategy of Conus. Exploration of this group of venom components offers an exciting new avenue for the discovery of novel pharmacological tools and drug candidates, complementary to conotoxins.


Asunto(s)
Péptidos/metabolismo , Caracoles , Ponzoñas/metabolismo , Animales , Conotoxinas
18.
Mar Drugs ; 15(5)2017 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-28531118

RESUMEN

The marine cone snail Conus gloriamaris is an iconic species. For over two centuries, its shell was one of the most prized and valuable natural history objects in the world. Today, cone snails have attracted attention for their remarkable venom components. Many conotoxins are proving valuable as research tools, drug leads, and drugs. In this article, we present the venom gland transcriptome of C. gloriamaris, revealing this species' conotoxin repertoire. More than 100 conotoxin sequences were identified, representing a valuable resource for future drug discovery efforts.


Asunto(s)
Conotoxinas/química , Conotoxinas/farmacología , Caracol Conus/fisiología , Venenos de Moluscos/química , Secuencia de Aminoácidos , Animales , Venenos de Moluscos/metabolismo , Transcriptoma
19.
Mol Cell Proteomics ; 13(4): 938-53, 2014 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-24478445

RESUMEN

Cone snails are highly successful marine predators that use complex venoms to capture prey. At any given time, hundreds of toxins (conotoxins) are synthesized in the secretory epithelial cells of the venom gland, a long and convoluted organ that can measure 4 times the length of the snail's body. In recent years a number of studies have begun to unveil the transcriptomic, proteomic and peptidomic complexity of the venom and venom glands of a number of cone snail species. By using a combination of DIGE, bottom-up proteomics and next-generation transcriptome sequencing the present study identifies proteins involved in envenomation and conotoxin maturation, significantly extending the repertoire of known (poly)peptides expressed in the venom gland of these remarkable animals. We interrogate the molecular and proteomic composition of different sections of the venom glands of 3 specimens of the fish hunter Conus geographus and demonstrate regional variations in gene expression and protein abundance. DIGE analysis identified 1204 gel spots of which 157 showed significant regional differences in abundance as determined by biological variation analysis. Proteomic interrogation identified 342 unique proteins including those that exhibited greatest fold change. The majority of these proteins also exhibited significant changes in their mRNA expression levels validating the reliability of the experimental approach. Transcriptome sequencing further revealed a yet unknown genetic diversity of several venom gland components. Interestingly, abundant proteins that potentially form part of the injected venom mixture, such as echotoxins, phospholipase A2 and con-ikots-ikots, classified into distinct expression clusters with expression peaking in different parts of the gland. Our findings significantly enhance the known repertoire of venom gland polypeptides and provide molecular and biochemical evidence for the compartmentalization of this organ into distinct functional entities.


Asunto(s)
Conotoxinas/genética , Conotoxinas/metabolismo , Caracol Conus/genética , Caracol Conus/metabolismo , Secuencia de Aminoácidos , Animales , Caracol Conus/clasificación , Células Epiteliales/citología , Células Epiteliales/metabolismo , Perfilación de la Expresión Génica , Variación Genética , Secuenciación de Nucleótidos de Alto Rendimiento , Datos de Secuencia Molecular , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteómica , Reproducibilidad de los Resultados , Alineación de Secuencia
20.
J Proteome Res ; 14(2): 688-99, 2015 Feb 06.
Artículo en Inglés | MEDLINE | ID: mdl-25412008

RESUMEN

The complex interplay of many cell types and the temporal heterogeneity of pancreatic islet composition obscure the direct role of resident alpha and beta cells in the development of Type 1 diabetes. Therefore, in addition to studying islets isolated from non-obese diabetic mice, we analyzed homogeneous cell populations of murine alpha (αTC-1) and beta (NIT-1) cell lines to understand the role and differential survival of these two predominant islet cell populations. A total of 56 proteins in NIT-1 cells and 50 in αTC-1 cells were differentially expressed when exposed to proinflammatory cytokines. The major difference in the protein expression between cytokine-treated NIT-1 and αTC-1 cells was free radical scavenging enzymes. A similar observation was made in cytokine-treated whole islets, where a comprehensive analysis of subcellular fractions revealed that 438 unique proteins were differentially expressed under inflammatory conditions. Our data indicate that beta cells are relatively susceptible to ER and oxidative stress and reveal key pathways that are dysregulated in beta cells during cytokine exposure. Additionally, in the islets, inflammation also leads to enhanced antigen presentation, which completes a three-way insult on beta cells, rendering them targets of infiltrating T lymphocytes.


Asunto(s)
Diabetes Mellitus Experimental/metabolismo , Retículo Endoplásmico/metabolismo , Islotes Pancreáticos/metabolismo , Estrés Oxidativo , Animales , Western Blotting , Islotes Pancreáticos/patología , Ratones , Ratones Endogámicos NOD
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