Rapid and efficient identification of cysteine-rich peptides by random screening of a venom gland cDNA library from the hexathelid spider Macrothele gigas

Satake, Honoo; Villegas, Elba; Oshiro, Naomasa; Terada, Koki; Shinada, Tetsuro

doi:10.1016/j.toxicon.2004.05.012

Cited by 28 publications

(19 citation statements)

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“…Several communications have been reported combinatorial strategies for exploring the transcriptome and the expressed venom components on different spider species. For example, the random screening of ESTs has been combined with proteomic approaches in spider transcriptome analyses of M. gigas (Satake et al 2004), H. schmidti or O. huwena Jiang et al 2010), C. crawshayi (Diego-Garcia et al 2010), H. hainanum or O. hainana , A. ventricosus , T. loki (Undheim et al 2013), and P. tristis (Zobel-Thropp et al 2014). In some cases, the proteomic approach showed a high number of peptide molecular masses compared to the obtained number of toxin-like transcripts (e.g., H. schmidti, Jiang et al 2008a;Jiang et al 2010).…”

Section: Comparative Data In Spider Venom Gland Transcriptomesmentioning

confidence: 99%

“…In 2001, the group of Byung Jin in Korea reported their results from the random screening of a cDNA library of the whole body from Araneus ventricosus (Chung et al 2001). Later, in 2004, the identification of cysteine-rich peptides by random screening of venom gland cDNA library from Macrothele gigas was used to obtain Magi toxins by the research group of Gerardo Corzo and H. Stake from Japan (Satake et al 2004). However, it was not until 2005 when the venom gland transcriptome analysis of the spider Agelena orientalis was published (Kozlov et al 2005).…”

Section: Introductionmentioning

confidence: 99%

“…(I) Transcriptomes from venom glands using cDNA cloning and Sanger sequencing approach were described from Macrothele gigas (Satake et al 2004) and Agelena orientalis (Kozlov et al 2005). The Brazilian group of Denise Tambourgi reported Loxosceles laeta (Fernandes-Pedrosa et al 2008) and by the group of Silvio Sanches L. intermedia (Gremski et al 2010).…”

Section: Introductionmentioning

confidence: 99%

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Spider Transcriptomes from Venom Glands: Molecular Diversity of Ion Channel Toxins and Antimicrobial Peptide Transcripts

et al. 2016

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Section: Comparative Data In Spider Venom Gland Transcriptomesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Spider Transcriptomes from Venom Glands: Molecular Diversity of Ion Channel Toxins and Antimicrobial Peptide Transcripts

et al. 2016

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“…Six peptide toxins (Magi 1-6) were isolated from the Hexathelidae spider, Macrothele gigas, dealing with sodium channel (Magi 1-5) and high toxicity in lepidoptera larvae of 3.1 nmol/g (Magi 6) [17] . Toxicological assays showed diverse lethal or paralytic activities of Magi 7-16 in mice and/or insects [18] . The spider Macrothele raveni was indentified as a new species in the genus Macrothele [19] .…”

Section: Introductionmentioning

confidence: 99%

Effects of spider Macrothele raven venom on cell proliferation and cytotoxicity in HeLa cells1

Gao

Shan

Chen

et al. 2005

Acta Pharmacologica Sinica

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Aim: To examine the effect of venom from the spider Macrothele raven on cell proliferation and cytotoxicity in human cervical carcinoma, HeLa cells. Methods: Morphological and biochemical signs of apoptosis appeared using acridine orange-ethidium bromide (AO/EB) staining. Marked morphological changes in HeLa cells after treatment with spider venom were observed using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Cell proliferation and cytotoxicity were determined by [methyl-3 H] thymidine assay ([ 3 H]TdR) and lactate dehydrogenase (LDH) release, respectively. DNA fragmentation and cell cycle distribution were monitored using flow cytometry. In addition, Western blot analysis was used to evaluate the level of caspase-3 expression. In vivo examination of the inhibition of the size of tumors in nude mice treated with spider venom was measured. Results: Marked morphological changes were observed using AO/EB staining, SEM and TEM assay. Spider venom at concentrations of 10-40 mg/L caused dose-and time-dependent inhibition of HeLa cell proliferation. The ratio of apoptosis and necrosis increased. The activity of caspase-3 was upregulated after spider venom treatment. In vivo study of tumor size revealed that tumors significantly decreased in size from controls to tumors treated for 3 weeks with spider venom (P<0.05). Conclusion: The inhibition of HeLa cells by the venom of the spider Macrothele raveni was carried out in three ways: induction of apoptosis, necrosis of toxicity damage and direct lysis. Spider venom is a novel anti-tumor material both in vitro and in vivo.

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“…Intra-thoracic injection of U 12 -HXTX-Mg1a (Magi-11) in crickets (Gryllus bimaculatus) induced slow-onset paralysis leading to death, while intracranial injection in mice resulted in lacrimation, sweating, and urination followed by paralysis of breathing and death 15 min post-injection. b-HXTX-Mg1a/b-HXTXMr1a caused reversible paralysis when injected into caterpillars (Spodoptera litura) and had no effect when injected into cockroaches (Periplaneta americana), whereas intraperitoneal injection in mice resulted in excitation, gasping, spastic paralysis, and exophthalmos (bulging of the eyes) followed by death (Satake et al, 2004). b-HXTXMg1a binds to site 4 on vertebrate Na V channels (see Fig.…”

Section: Nasptx Familymentioning

confidence: 99%

Developing novel analgesics to treat chronic pain: inhibiting voltage gated sodium channel 1.7 with spider-venom peptides

Arachchilage¹

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Chronic pain is responsible for great physical, mental and economic loss. While there are diverse methods available for managing pain, such as NSAID, opioids and local anesthetics, common issues associated such as addiction and tolerance highlights the unmet demand for a novel analgesic aimed at a novel target.Numerous reports on individuals suffering from loss of function mutations in SCN9A marker, which leads to complete inability to feel pain, yet otherwise normal physiology, has led to the recognition of voltage gated sodium channel 1.7 (Na V 1.7) as a prime pain target.Other reports of gain of function mutations that lead to conditions such as paroxysmal extreme pain disorder and primary erythromelalgia confirms the validity of this target.However the presence of nine closely related subtypes of voltage gated sodium channels, Na V 1.1-Na V 1.9 introduces complications as any cross inhibition may lead to detrimental results. For example inhibition of Na V 1.5 subtype could result in cardiac arrhythmia to complete cardiac arrest and death. Therefor it is of utmost importance that any pharmaceutical agent used to inhibit Na V 1.7 is highly specific for Na V 1.7 and display no cross reactivity on other voltage gated channels.Spiders are one of the most successful terrestrial predators; their venoms have evolved over many millions of years to selectively and potently inhibit nervous system targets in order to rapidly paralyze their prey. Spider-venom peptidomes have been identified as one of the largest known libraries of compounds with very high potency and specificity towards nervous system targets.Spider toxins listed in the Arachnoserver database have been classified into 12 families of sodium channel toxins (NaSpTx), denoted NaSpTx Families 1-12, based on the level of sequence conservation and intercystine spacing. NaSpTx family 2 is the largest toxin family; it comprises 34 peptides of theraphosid origin.This thesis primarily aims at discovering novel inhibitors of Na V 1.7. However the intriguing fact that people suffering form congenital indifference to pain also suffers form anosmia, has lead to efforts of understanding the role of Na V 1.7 in olfaction. This study has been iii completed with the finding that Na V 1.7 is located along axons of olfactory neurones. These findings have been published and research article is attached as chapter 4.During the initial discovery phase of this project, an assay guided fractionation method was used to identify Na V 1.7 inhibitors form six spider venoms. This resulted in nine fully sequenced peptides where four of these belonged to NaSpTx family 2. This included the well-known β/ω-TRTX-Tp1a (Protoxin1). Three novel peptides discovered were named β-TRTX-Pe1a, µ-TRTX-Pe1b, and U-TRTX-Pa1a.A complete alanine scan of β/ω-TRTX-Tp1a on Na V 1.7 was conducted using a Xenopus laevis oocyte based tethered toxin method. Residues that were important for Na V 1.7 binding were mapped onto the three-dimensional structure of β/ω-TRTX-Tp1a. Three novel toxin peptides β-TR...

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Rapid and efficient identification of cysteine-rich peptides by random screening of a venom gland cDNA library from the hexathelid spider Macrothele gigas

Cited by 28 publications

References 31 publications

Spider Transcriptomes from Venom Glands: Molecular Diversity of Ion Channel Toxins and Antimicrobial Peptide Transcripts

Spider Transcriptomes from Venom Glands: Molecular Diversity of Ion Channel Toxins and Antimicrobial Peptide Transcripts

Effects of spider Macrothele raven venom on cell proliferation and cytotoxicity in HeLa cells1

Developing novel analgesics to treat chronic pain: inhibiting voltage gated sodium channel 1.7 with spider-venom peptides

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