Cloning and characterization of an acidic cytolysin cDNA from sea anemone Sagartia rosea

Jiang, Xiao; Wen, Yunfei; Chen, Hui Ping; Tu, Hong Bin; Wu, Wen Yan; Wei, Jian Wen; Wang, Juan; Liu, Wenhua; An, Xiaobo

doi:10.1016/s0041-0101(02)00173-3

Cited by 23 publications

(8 citation statements)

References 20 publications

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“…Previous actinoporin gene tree reconstructions formed taxonomically distinct groupings, all of which were recovered to some degree in our analysis. However, a large, strongly supported gene cluster containing sequences from actinioideans (see Figure 4, [1]), now includes several non-actinioidean taxa: Sagartia elegans (see [62]), Metridium senile , Andvakia discipulorum , Calliactis parasitica , and Stomphia coccinea (Figure 2). Sister to this gene cluster is a strongly supported gene cluster of newly identified candidate actinoporins, determined from primarily genomic data from Heteractis crispa and Epiactis japonica (Figure 2).…”

Section: Discussionmentioning

confidence: 99%

Evolution of the Cytolytic Pore-Forming Proteins (Actinoporins) in Sea Anemones

Macrander

Daly

2016

Toxins

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Sea anemones (Cnidaria, Anthozoa, and Actiniaria) use toxic peptides to incapacitate and immobilize prey and to deter potential predators. Their toxin arsenal is complex, targeting a variety of functionally important protein complexes and macromolecules involved in cellular homeostasis. Among these, actinoporins are one of the better characterized toxins; these venom proteins form a pore in cellular membranes containing sphingomyelin. We used a combined bioinformatic and phylogenetic approach to investigate how actinoporins have evolved across three superfamilies of sea anemones (Actinioidea, Metridioidea, and Actinostoloidea). Our analysis identified 90 candidate actinoporins across 20 species. We also found clusters of six actinoporin-like genes in five species of sea anemone (Nematostella vectensis, Stomphia coccinea, Epiactis japonica, Heteractis crispa, and Diadumene leucolena); these actinoporin-like sequences resembled actinoporins but have a higher sequence similarity with toxins from fungi, cone snails, and Hydra. Comparative analysis of the candidate actinoporins highlighted variable and conserved regions within actinoporins that may pertain to functional variation. Although multiple residues are involved in initiating sphingomyelin recognition and membrane binding, there is a high rate of replacement for a specific tryptophan with leucine (W112L) and other hydrophobic residues. Residues thought to be involved with oligomerization were variable, while those forming the phosphocholine (POC) binding site and the N-terminal region involved with cell membrane penetration were highly conserved.

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Section: Discussionmentioning

confidence: 99%

Evolution of the Cytolytic Pore-Forming Proteins (Actinoporins) in Sea Anemones

Macrander

Daly

2016

Toxins

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“…An arrow denotes the thrombin cleavage site. (D) The alignment of known N-terminal sequences of actinoporins compiled from the literature [8,[45][46][47][48][49]. Hydrophobic amino acids (AGVLIFWYP) are shown in black, polar amino acids (TSMCNQ) are shown in dark gray, and charged amino acids (DEKHR) are shown in light gray.…”

Section: Characterization Of Transmembrane Channels Formed By Eqtii mentioning

confidence: 99%

The equinatoxin N‐terminus is transferred across planar lipid membranes and helps to stabilize the transmembrane pore

et al. 2006

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Equinatoxin II is a cytolytic protein isolated from the sea anemone Actinia equina. It is a member of the actinoporins, a family of eukaryotic pore‐forming toxins with a unique mechanism of pore formation. Equinatoxin II is a 20 kDa cysteineless protein, with sphingomyelin‐dependent activity. Recent studies showed that the N‐terminal region of the molecule requires conformational flexibility during pore formation. An understanding of the N‐terminal position in the final pore and its role in membrane insertion and pore stability is essential to define the precise molecular mechanism of pore formation. The formation of pores and their electrophysiologic characteristics were studied with planar lipid membranes. We show that amino acids at positions 1 and 3 of equinatoxin II are exposed to the lumen of the pore. Moreover, sulfhydryl reagents and a hexa‐histidine tag attached to the N‐terminus revealed that the N‐terminus of the toxin extends through the pore to the other (trans) side of the membrane and that negatively charged residues inside the pore are crucial to define the electrophysiologic characteristics of the channel. Finally, we detected a new, less stable, state with a lower conductance by using a deletion mutant in which the first five N‐terminal amino acids were removed. We propose that the first five amino acids help to anchor the amphipathic helix on the trans side of the membrane and consequently stabilize the final transmembrane pore.

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“…Different studies comparing the genes structure of actinoporins from genomic samples and cDNA revealed that the genes lack introns within the coding region. The lack of introns in actinoporins have been reported in Eqts (Anderluh et al, 1996;Punger car et al, 1997), SrcI (Jiang et al, 2002), and HMgs .…”

Section: Genetic Structure Of Actinoporinsmentioning

confidence: 75%

“…The regions of translation start of the mature sequences of actinoporins include the base adenine (A) highly conserved at Kozak consensus sequence at positions À3 and þ4 (Kozak, 1987) and to the 3 0 -end of the translated sequences also contain a putative polyadenylation signal (a typical polyA tail eukaryotic signal sequence) (Anderluh et al, 1996;Jiang et al, 2002). Different studies comparing the genes structure of actinoporins from genomic samples and cDNA revealed that the genes lack introns within the coding region.…”

Section: Genetic Structure Of Actinoporinsmentioning

confidence: 99%

“…Actinoporins are small proteins (~20 kDa) with only one domain and are classified as PFP a-type due to their mode of membrane integration through a-helical elements AlegreCebollada et al, 2007a). Their isoelectric point (pI) is generally basic, usually above nine, with exception to the epiactin-A from Epiactis prolifera (pI ¼ 6.4) and SrcI from Sagartia rosea (pI ¼ 4.8), which have been described as acidic actinoporins (Bernheimer and Avigad, 1982;Jiang et al, 2002). Another important characteristic of actinoporins is their inhibition by preincubation with sphingomyelin (SM), which led to the proposal that this lipid could be the membrane target Alegre-Cebollada et al, 2007a).…”

Section: Introductionmentioning

confidence: 99%

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