MINIREVIEW
Marine toxins and the cytoskeleton: pectenotoxins,
unusual macrolides that disrupt actin
Begon
˜
a Espin
˜
a
1
and Juan A. Rubiolo
1,2
1 Departamento de Farmacologia, Facultad de Veterinaria, Universidad de Santiago de Compostela, Lugo, Spain
2 Departamento de Fisiologia Animal, Facultad de Veterinaria, Universidad de Santiago de Compostela, Lugo, Spain
The pectenotoxins (PTXs), macrolactones with multi-
ple polyether ring units that have been shown to con-
taminate shellfish in various parts of the world (Fig. 1)
[1–7], were first isolated from the Japanese scallop
Patinopecten yessoensis [8].
PTX-2 is produced by many species of the dinofla-
gellate genus Dinophysis, and it was initially detected
in Dinophysis fortii [9]. Later, this toxin was isolated
from Dinophysis acuminate, Dinophysis norvegica, Din-
ophysis rotundata and Dinophysis acuta [1–3,6,10–13].
After consumption of the algae by the shellfish, PTX-2
can be metabolized to other PTX derivatives. In the
digestive gland of the scallop P. yessoensis, the C43
methyl group in PTX-2 is oxidized to the alcohol
(PTX-1), aldehyde (PTX-3) and carboxylic acid (PTX-
6) forms [7,13]. Also, PTX-4 and PTX-7 have been
isolated from the digestive glands of scallops collected
in Japan, and these are stereoisomers of PTX-1 and

Intraperitoneal injection of pectenotoxins into mice produces high hepato-
toxicity as the principal symptom, so the liver seems to be their target
organ. Up to now, 15 pectenotoxin analogs have been discovered, with dif-
ferent toxicological potencies that are related to their structures. Now, it is
generally accepted that the actin cytoskeleton is the principal molecular
target of pectenotoxins. Although recent studies have demonstrated that
pectenotoxins induce actin filament disruption by a capping effect, other
kinds of activity, such as sequestration of actin, cannot be ruled out. All of
the active analogs tested triggered disruption of the actin cytoskeleton and
displayed potencies that correlated with their toxicity in mice. Moreover,
pectenotoxins induce apoptosis to a higher degree in tumor cells than in
normal cells of the same tissue. This fact opens the prospect of studying
new chemotherapy agents and actin cytoskeleton dynamics with potential
clinical applications.
Abbreviations
F-actin, filamentous actin; G-actin, globular actin; OA, okadaic acid; PTX, pectenotoxin; SA, seco acid.
6082 FEBS Journal 275 (2008) 6082–6088 ª 2008 The Authors Journal compilation ª 2008 FEBS
the seco acid form. Epimerization of PTX-2 SA yields
the thermodynamically more stable 7-epi-PTX-2 SA,
and both have been detected in shellfish from Portugal
[6], Ireland [2], New Zealand [4] and Croatia [3].
Besides PTX-2, four other oxidized forms of this
toxin, PTX-11, PTX-12, PTX-13 and PTX-14, have
been isolated from samples of Dinophysis [12,15–17].
PTX-11 is the 34S-hydroxy PTX-2 and is more resis-
tant to enzymatic hydrolysis than PTX-2 when
exposed to mussel hepatopancreas [17]. The PTX-
12 SA ⁄ PTX-12 ratio in naturally contaminated mussel
samples is markedly lower than the PTX-2 SA ⁄ PTX-2
ratio [12], suggesting that PTX-12 is more resistant to

route than via the intraperitoneal route. It is now
accepted that the PTXs do not induce diarrhea [24,25].
PTXs are toxic to the liver when administered intra-
peritoneally in mice, PTX-2 being the most potent.
Oxidation of this toxin to PTX-1, PTX-3 and PTX-6 is
accompanied by a decrease in toxicity [7]. PTX-11 has
been reported to be as toxic as PTX-2, producing the
same symptoms of intoxication in mice [17]. Ito et al.
reported that the liver injuries produced by PTX-6 are
different from those produced by PTX-2; whereas
PTX-2 produced congestion under the liver capsule as
a result of circulatory disorder, PTX-6 caused severe
bleeding in the liver [26].
The first to establish the hepatotoxicity of these
toxins were Terao et al., who showed that PTX-1
produced liver damage after intraperitoneal injection
into mice, inducing necrosis of hepatocytes, principally
in the periportal regions of the hepatic lobules [25].
Hepatocyte death was also observed by Fladmark
et al. [27] in freshly isolated rat and salmon hepato-
cytes, but in this case PTX-1 induced apoptosis rather
than necrosis according to the chromatin hyperconden-
sation, cell shrinkage and lack of Trypan blue uptake
observed [27].
It has been shown that PTX-1, PTX-2, PTX-6 and
PTX-11 disrupt the filamentous actin (F-actin)
cytoskeleton in NRK-52E cells, rabbit enterocytes and
neuroblastoma cells [28–31], and it is proposed that
PTXs exert their toxicity by this mechanism.
PTX structure–activity relationship

nontoxic to mice by the oral and intraperitoneal routes
at 5000 lgÆkg
)1
[23].
Toxicological studies indicated evident differences
between PTX potencies that can be related to their
particular structures (Fig. 1). Analogs isomerized so as
to contain a six-membered B-ring, PTX-8 and PTX-9,
are more than one order of magnitude less toxic than
those containing a five-membered B-ring. They are
believed to be artefacts produced during the purifica-
tion process [14]. PTX-3 and PTX-6, the 7R-epimers
of PTX-4 and PTX-7, are significantly less toxic than
their corresponding 7S-epimers [7].
Only two in vitro studies have compared the effects
of diverse PTXs on cells, and both agree with the
above cited in vivo toxicological assays. Daiguji et al.
reported that PTX-2 SA was not toxic to KB cells
even at 1.8 lgÆmL
)1
, whereas PTX-2 was cytotoxic at
0.05 lgÆmL
)1
[10]. Ares et al. found that PTX-1,
PTX-2 and PTX-11 modified the actin cytoskeleton
and morphology of BE(2)-M17 neuroblastoma cells.
However, PTX-2 SA did not show any effect on these
cells. Moreover, PTX-2 and PTX-11 were more
potent than PTX-1 (Fig. 2) [28]. This supports the
idea that lactone ring integrity is essential for the

the halichondramides, exhibit barbed-end capping and
F-actin-severing activity. Moreover, unusual dimeric
macrolides, swinholide-A and misakinolide-A, were,
surprisingly, found to display different effects on actin
dynamics; swinkolide-A severs actin, whereas misakino-
lide-A rapidly caps barbed ends of filaments [31].
The actin cytoskeleton – molecular
target of PTXs
Initial studies performed to elucidate the mechanism of
action of PTXs and their in vitro biochemical effects
demonstrated that PTX-2 inhibited actin polymeriza-
tion in a concentration-dependent manner [36]. PTX-2
also inhibited contractions induced by KCl in the
isolated rat aorta, and formed a 1 : 4 complex with
G-actin [36,37]. Spector et al. reported that PTX-2
sequestered monomeric actin with a K
d
of 20 nm, but
did not exhibit severing or capping activity. Moreover,
PTX-2 disrupted the organization of actin in several
cell types in a time- and concentration-dependent
manner [31].
In a recent study, Allingham et al. obtained the
X-ray structure of the PTX-2–actin complex. In con-
trast to the results of Hori et al., they described a 1 : 1
stoichiometry in a novel site between subdomains 1
and 3 of the actin molecule [38]. In cells, capping pro-
teins such as gelsolin play an important role; capping
of old filaments funnels actin polymerization to sites
where new barbed ends are present, and the generation

Cytological implications of disruption
of the actin cytoskeleton
Although, according to the histopathological studies,
the liver seems to be the target organ of PTXs, many
cellular models have shown substantial effects of PTX
treatments in the nanomolar range [27,40–42]. The
induction of apoptotic cell death rather than necrosis
by PTX-1 and PTX-2 is well established, on the basis
of morphological changes, reduction of mitochondrial
membrane potential, increases in cytoplasmic cyto-
chrome c and Smac ⁄ DIABLO, and caspase-3 and
caspase-9 activation. The inhibition of PTX-induced
cell death by caspase inhibitors supports the apoptotic
pathway [27,40].
Actin polymerization and dynamics are required for
essential cell processes such as motility, endocytosis,
cytokinesis, and establishing cell–cell and cell–substrate
contacts. In vitro and in vivo studies showed that some
macrolide–actin complexes reduced free G-actin below
the critical concentration and blocked extension of the
(+)-end of the filament, while the free macrolide also
bound to and severed actin filaments [43]. In living
cells, the combined effects of these interactions leads
to a cessation of new filament growth and a disruption
of existing filaments, and is accompanied by a loss of
motility, breakdown of adherens junctions, polyploidy
and, ultimately, apoptosis [43].
Very few studies have compared the effects of PTXs
in diverse cellular models. Chae et al. found that PTX-2
induced apoptosis in p53-deficient cells [40], showing

DNAse I, and visualized with a confocal
microscope. Scale bar: 50 lm.
Actin cytoskeleton disruption by pectenotoxins B. Espin˜ a and J. A. Rubiolo
6086 FEBS Journal 275 (2008) 6082–6088 ª 2008 The Authors Journal compilation ª 2008 FEBS
Perspectives
The importance of the actin cytoskeleton in pathogenic
cellular process such as angiogenesis, cell adhesion,
cytokinesis and metastasis has made it an attractive
target for the development of anticancer drugs. Drugs
that block the regulation of actin filament dynamics
within tumor cells or in cells infected with pathogens
could be useful in treating cancer and other diseases.
In addition, differential actions of antiactin com-
pounds can reveal interesting data about actin cyto-
skeleton dynamics.
Although the presence in the food chain of PTXs is
being regulated, together with that of diarrhetic shell-
fish poisoning toxins, toxicological tests by oral admin-
istration point to these compounds as nonhazardous
molecules for human health with regard to shellfish
consumption. However, the selective cytotoxicity of
PTXs against certain tumor cell lines and their ability
to induce apoptosis of p53-deficient cell lines, which are
often resistant to other chemotherapeutic agents, makes
them interesting new candidates for cancer therapy.
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