Elsevier

Peptides

Volume 47, September 2013, Pages 71-76
Peptides

Functional characterization on invertebrate and vertebrate tissues of tachykinin peptides from octopus venoms

https://doi.org/10.1016/j.peptides.2013.07.002Get rights and content

Highlights

  • Octopus venoms contain tachykinin peptides.

  • These peptides are more similar to vertebrate non-venom types than invertebrate.

  • These peptides are active on both invertebrate and vertebrate receptors.

Abstract

It has been previously shown that octopus venoms contain novel tachykinin peptides that despite being isolated from an invertebrate, contain the motifs characteristic of vertebrate tachykinin peptides rather than being more like conventional invertebrate tachykinin peptides. Therefore, in this study we examined the effect of three variants of octopus venom tachykinin peptides on invertebrate and vertebrate tissues. While there were differential potencies between the three peptides, their relative effects were uniquely consistent between invertebrate and vertebrae tissue assays. The most potent form (OCT-TK-III) was not only the most anionically charged but also was the most structurally stable. These results not only reveal that the interaction of tachykinin peptides is more complex than previous structure–function theories envisioned, but also reinforce the fundamental premise that animal venoms are rich resources of novel bioactive molecules, which are useful investigational ligands and some of which may be useful as lead compounds for drug design and development.

Introduction

Tachykinins are a highly conserved group of peptides found in both invertebrate and vertebrate animals. These peptides function as neurotransmitters and neuromodulators of both the central and peripheral nervous systems [55]. The mammalian tachykinins neurokinin A, neurokinin B and Substance P are sensory neuropeptides with roles in both nociception and inflammation. Tachykinins exhibit both afferent and efferent functions and participate in the regulation of several physiological processes including peripheral sensory mechanisms such as nociception and inflammation as well as autonomic functions such as smooth muscle contractility in the vascular, gastrointestinal and genitourinary systems [34], [44]. In addition, tachykinins are involved in central nervous system pathways mediating pain, anxiety, motor coordination and cognition [34].

The actions of tachykinin peptides are mediated by one or more tachykinin receptors. Three subtypes of vertebrate tachykinin receptors, known as neurokinin receptor 1 (NK1R), neurokinin receptor 2 (NK2R), and neurokinin receptor 3 (NK3R), as well as numerous subtypes of invertebrate tachykinin receptors, have been described to date [37], [55]. Neurokinin receptors have been shown to act via Gq/11 coupling proteins increasing inositol phosphate 3 and diaceylglycerol (DAG) levels within cells bound by an agonist [41]. To date, a number of characteristic tachykinin amino acid motifs have been found to be crucial to the structure–activity relationships of tachykinins and tachykinin-like peptides. Vertebrate tachykinins are characterized by a FXGLM-amide motif while invertebrate tachykinins are characterized by a C-terminal FXGXR-amide motif (Table 1).

Octopuses live in habitats ranging from pelagic to benthic zones of all of the world's oceans ranging from Arctic to Antarctic, with some species specialists to certain habitats [53]. Octopuses secrete a variety of bioactive molecules from their posterior venom glands in order to feed on both vertebrate and invertebrate prey [15], [24], [26]. Upon envenomation rapid immobilization due to hypotensive effects as well as complete, irreversible, flaccid paralysis is observed in crustaceans [21], [40]. The consequent evolutionary selection pressure has resulted in a wide diversity of bioactive substances present in octopus and other coleoid venoms including small molecules such as acetylcholine, histamine, octopamine, tserotonin (aka: enteramine), taurine, tetrodotoxin and tyramine [7], [8], [10], [11], [12], [13], [14], [16], [17], [20], [29], [47] and proteins [1], [3], [18], [19], [22], [23], [25], [33], [36], [38], [45], [48], [51], [52]. Included in this tremendous molecular biodiversity are the tachykinin peptides Oct-TK-I and Oct-TK-II from Octopus vulgaris [32], Oct-TK-III from Octopus kaurna [19], and eledoisin from Eledone cirrhosa [1], [23]. These tachykinin forms are interesting in that even though they are from an invertebrate venom, the C-terminal amide motif is that of the vertebrate form (Table 2).

As octopuses prey upon a wide diversity of vertebrate and invertebrate species, one might predict their tachykinin type toxins to target the vertebrate as well as the invertebrate receptors but this prediction has never been experimentally tested. Structurally, only vertebrate-type tachykinins containing a C-terminal FXGLM-amide moiety have thus far been identified in octopus venom. Consistent with this observation, the tachykinins Oct-TK-I and Oct-TK-II from octopus venom have been shown to be vertebrate active [32]. However a comparison of the relative effects upon vertebrate and vertebrate tissues has not been undertaken. Further, a novel tachykinin we previously sequenced [19], which differs significantly from Oct-TK-I and Oct-TK-II in nature and distribution of charged residues, has not been functionally characterized. Therefore, the aim of this study was to compare the differential effects of octopus venom tachykinin peptides upon invertebrate and vertebrate tissue preparations.

Section snippets

Peptide synthesis and purification

In order to explore potential neofunctionalization derivations, we constructed Oct-TK-I (KPPSSSEFIGLM), Oct-TK-II (KPPSSSEFVGLM) and Oct-TK-III (DPPSDDEFVSLM) peptides in both amide and non-amide forms. Protected Fmoc-amino acid derivatives were purchased from Auspep (Melbourne, Australia). The following side chain protected amino acids were used: His(Trt), Hyp(tBu), Tyr(tBu), Lys(Boc), Trp(Boc), Arg(Pbf), Asn(Trt), Asp(OtBu), Glu(OtBu), Gln(Trt), Ser(tBu), Thr(tBu), and Tyr(tBu). All other

Results

In the vertebrate-isolated ileum tissue assay, all three amide-peptides showed classic tachykinin responses: after addition to tissue there was an initial decrease in contractility, followed by an increase to peak concentration-dependent contraction, and then a period of oscillation until the peptide was washed from the organ bath (Fig. 1, Fig. 2). When tested on the invertebrate hindgut assay, all three peptides elicited increases in contraction (Fig. 3), with similar EC50 values (Table 2).

Discussion

In this study, we examined the differential effects of the octopus venom peptides Oct-TK-I, Oct-TK-II and Oct-TK-III on vertebrate and invertebrate tissue-specific contractile activity using the rat ileum (Rattus norvegicus) and crayfish hindgut (P. clarkia) assays. Our findings indicate that OCT-TK-I, OCT-TK-II and OCT-TK-III are differentially active when assayed for invertebrate- and vertebrate-specific effects. For all three peptides, the C-terminal amide was essential for any activity,

Acknowledgements

BGF was funded by an Australian Research Council Future Fellowship and by the University of Queensland. SAA was the recipient of postdoctoral fellowship (PDRF Phase II Batch-V) from Higher Education Commission (HEC Islamabad) Pakistan. TNWJ was funded by an Australian Postgraduate Award. EABU acknowledges funding from the University of Queensland (International Postgraduate Research Scholarship, UQ Centennial Scholarship, and UQ Advantage Top-Up Scholarship) and the Norwegian State Education

References (58)

  • T. Ikeda et al.

    The importance of C-terminal residues of vertebrate and invertebrate tachykinins for their contractile activities in gut tissues

    FEBS Lett

    (1999)
  • M.W. Jarvis et al.

    Chromatographic properties of maculotoxin, a toxin secreted by Octopus (Hapalochlaena) maculosus

    Toxicon

    (1975)
  • E.C. Johnson et al.

    Identification of Drosophila neuropeptide receptors by G protein-coupled receptors-beta-arrestin2 interactions

    J Biol Chem

    (2003)
  • A. Kanda et al.

    Isolation and characterization of novel tachykinins from the posterior salivary gland of the common octopus Octopus vulgaris

    Peptides

    (2003)
  • A.M. Khawaja et al.

    Tachykinins: receptor to effector

    Int J Biochem Cell Biol

    (1996)
  • C.A. Maggi

    The mammalian tachykinin receptors

    Gen Pharmacol

    (1995)
  • R.D. Pinnock et al.

    Characterization of tachykinin mediated increases in [Ca2+]i in Chinese hamster ovary cells expressing human tachykinin NK3 receptors

    Eur J Pharmacol

    (1994)
  • J. Poels et al.

    Characterization and distribution of NKD, a receptor for Drosophila tachykinin-related peptide 6

    Peptides

    (2009)
  • J. Poels et al.

    Functional comparison of two evolutionary conserved insect neurokinin-like receptors

    Peptides

    (2007)
  • H. Torfs et al.

    Analysis of C-terminally substituted tachykinin-like peptide agonists by means of aequorin-based luminescent assays for human and insect neurokinin receptors

    Biochem Pharmacol

    (2002)
  • A. Ueda et al.

    Purification and molecular cloning of SE-cephalotoxin, a novel proteinaceous toxin from the posterior salivary gland of cuttlefish Sepia esculenta

    Toxicon

    (2008)
  • E.A. Undheim et al.

    Venom on ice: first insights into Antarctic octopus venoms

    Toxicon

    (2010)
  • E.A. Undheim et al.

    Genetic identification of Southern Ocean octopod samples using mtCOI

    C R Biol

    (2010)
  • T. Van Loy et al.

    Tachykinin-related peptides and their receptors in invertebrates: a current view

    Peptides

    (2010)
  • A. Anastasi et al.

    Occurrence and some properties of eledoisin in extracts of posterior salivary glands of Eledone

    Br J Pharmacol Chemother

    (1962)
  • R.T. Birse et al.

    Widely distributed Drosophila G-protein-coupled receptor (CG7887) is activated by endogenous tachykinin-related peptides

    J Neurobiol

    (2006)
  • D.E. Champagne et al.

    Sialokinin I and II: vasodilatory tachykinins from the yellow fever mosquito Aedes aegypti

    Proc Natl Acad Sci USA

    (1994)
  • A.E. Christie et al.

    Two novel tachykinin-related peptides from the nervous system of the crab Cancer borealis

    J Exp Biol

    (1997)
  • H. Dircksen et al.

    Genomics, transcriptomics, and peptidomics of Daphnia pulex neuropeptides and protein hormones

    J Proteome Res

    (2011)
  • Cited by (18)

    • Pharmacological importance of TG12 from tachykinin and its toxicological behavior against multidrug-resistant bacteria Klebsiella pneumonia

      2021, Comparative Biochemistry and Physiology Part - C: Toxicology and Pharmacology
      Citation Excerpt :

      The tachykinin molecular function was mediated by tachykinin receptor; however, both vertebrate and invertebrate have different subtypes of receptors (Kumaresan et al., 2019). The vertebrate and invertebrate tachykinins were characterized by the presence of ‘FXGLM’ and ‘FXGXR’ amide motif at C-terminal, respectively (Kumaresan et al., 2019; Ruder et al., 2013). The first tachykinin was isolated from the intestine of an elasmobranch fish, Scyliorhinus canicula (Jiang et al., 2013).

    • The evolution and origin of tetrodotoxin acquisition in the blue-ringed octopus (genus Hapalochlaena)

      2019, Aquatic Toxicology
      Citation Excerpt :

      One prominent hypothesis proposes that proteinaceous venom evolution occurs through a process of duplication and neofunctionalization where by a copy of an innocuous key-processing gene is requisitioned for a new purpose (Casewell et al., 2013). Proteinaceous toxins form the lethal component of several octopod and cuttlefish venoms examined to date (Fry et al., 2009a; Ruder et al., 2013a, b; Undheim et al., 2010). Hapalochlaena represents an exception with the inclusion of the potent non-proteinaceous toxin TTX.

    • A combined proteomic and transcriptomic analysis of slime secreted by the southern bottletail squid, Sepiadarium austrinum (Cephalopoda)

      2016, Journal of Proteomics
      Citation Excerpt :

      Cephalotoxin homologues have also been found in a small number of other organisms [73,74] however its function is unknown in these species. Tachykinins are found in octopods [17] however within cuttlefish Cornet et al. [21] was unable to identify tachykinins within the PSG despite using tachykinin specific search methods. They were identified however, within the central nervous system as well as hemolymph, identifying other potential roles for tachykinins other than specific toxicity [75].

    • Dual role of the cuttlefish salivary proteome in defense and predation

      2014, Journal of Proteomics
      Citation Excerpt :

      Oct-TKI/II from Octopus vulgaris PSG has even been described as a homolog to substance P and is believed to induce death by cardiac and respiratory arrest [21]. Further, Oct-TKI/II/III exhibited activity on tissues of vertebrates and invertebrates that are both octopus preys [35]. Our search for tachykinin-like peptides based on the FXXLMamide pattern, using Peptraq software and based on peptidic extracts treated to contain only amidated peptides remained fruitless and led us to conclude that no tachykinin was produced in Sepia officinalis PSG.

    View all citing articles on Scopus
    1

    Joint first-authorship.

    View full text