Characterization of selectivity and pharmacophores of type 1 sea anemone toxins by screening seven Na-v sodium channel isoforms

During their evolution, animals have developed a set of cysteine-rich peptides capable of binding various extracellular sites of voltage-gated sodium channels (VGSC). Sea anemone toxins that target VGSCs delay their inactivation process, but little is known about their selectivities. Here we report the investigation of three native type 1 toxins (CGTX-II, delta-AITX-Bcg1a and delta-AITX-Bcg1b) purified from the venom of Bunodosoma cangicum. Both delta-AITX-Bcg1a and delta-AITX-Bcg1b toxins were fully sequenced. The three peptides were evaluated by patch-clamp technique among Nav1.1-1.7 isoforms expressed in mammalian cell lines, and their preferential targets are Na(v)1.5 > 1.6 > 1.1. We also evaluated the role of some supposedly critical residues in the toxins which would interact with the channels, and observed that some substitutions are not critical as expected. In addition, CGTX-II and delta-AITX-Bcg1a evoke different shifts in activation/inactivation Boltzmann curves in Nav1.1 and 1.6. Moreover, our results suggest that the interaction region between toxins and VGSCs is not restricted to the supposed site 3 (S3-54 linker of domain IV), and this may be a consequence of distinct surface of contact of each peptide vs. targeted channel. Our data suggest that the contact surfaces of each peptide may be related to their surface charges, as CGTX-II is more positive than delta-AITX-Bcg1a and delta-AITX-Bcg1b. (C) 2011 Elsevier Inc. All rights reserved.

Sobrevivência e crescimento inicial de Ocotea pulchella (Lauraceae) em uma floresta de restinga da Ilha do Cardoso, SP

Neste trabalho, avaliou-se os efeitos de fatores ambientais na sobrevivência e crescimento inicial de Ocotea pulchella (Nees) Mez numa floresta de restinga. Os ensaios consistiram do plantio de mudas classificadas em dois grupos de idade (plântulas e juvenis) em diferentes áreas selecionadas quanto à cobertura vegetal (clareiras ou sub-bosque) e umidade do solo (secos e úmidos). As plantas de ambas as idades apresentaram a maior taxa de mortalidade no sub-bosque úmido, onde se observou os valores mais baixos de luz fotossinteticamente ativa e fertilidade do solo. A sobrevivência de plântulas correlacionou-se positivamente ao crescimento, que por sua vez, foi favorecido em clareiras, sugerindo uma dependência da espécie em relação a ambientes mais iluminados, embora essa possa ocupar o sub-bosque em fisionomias mais abertas como as restingas. Em juvenis, a sobrevivência foi relacionada principalmente aos fatores edáficos; o crescimento em altura foi muito baixo, não sendo constatado incremento significativo de área foliar nem de matéria seca em nenhum dos ambientes, o que pode estar relacionado ao caráter oligotrófico do solo. Os juvenis em sub-bosque de solo úmido apresentaram ao final do experimento os menores valores de altura, área foliar, razão de área foliar, razão de massa foliar e taxa de assimilação líquida, sugerindo que baixas irradiâncias associadas a solos saturados e quimicamente pobres podem restringir o recrutamento da espécie no local estudado. Todavia, o comportamento de certa forma generalista para as condições de luz e umidade do solo, indica uma alta capacidade de O. pulchella em colonizar distintos micro-ambientes da restinga, onde se observa uma grande variação espaço-temporal de fatores ambientais.

CLEANER SHRIMP (CARIDEA: PALAEMONIDAE) ASSOCIATED WITH SCYPHOZOAN JELLYFISH

Shrimp are found in association with many hosts (sponges, anthozoans, mollusks, echinoids, ascidians and fish). Shrimp are rarely observed in association with jellyfish. The interaction between the cleaner shrimp Periclimenes paivai and jellyfish is poorly known, with only one published report for Brazilian waters. We add new information about this association, based on field and laboratory observations. Shrimp were collected from three locations on the coast Of Sao Paulo State (Cananeia, Santos and Sao Sebastiao) and from two species of scyphozoan rnedusae: Chrysaora lactea and Lychnorhiza lucerna. We classify the association as facultative commensalism, since the shrimp gain protection and probably feed on mucus produced by the jelly fish. The maximum carapace length of the shrimp collected was 9.2 min. Medusa bel I diameter ranged between about 10 and 19 cm for C. lactea and between 4.5 and 25 cm for L. lucerna. We also provide a list of records of Periclimenes in association with different scyphornedUsae, and additional information on the size and coloration of P. paivai.

Peptide fingerprinting of the neurotoxic fractions isolated from the secretions of sea anemones Stichodactyla helianthus and Bunodosoma granulifera. New members of the APETx-like family identified by a 454 pyrosequencing approach

Sea anemones are known to contain a wide diversity of biologically active peptides, mostly unexplored according to recent peptidomic and transcriptomic studies. In the present work, the neurotoxic fractions from the exudates of Stichodactyla helianthus and Bunodosoma granulifera were analyzed by reversed-phase chromatography and mass spectrometry. The first peptide fingerprints of these sea anemones were assessed, revealing the largest number of peptide components (156) so far found in sea anemone species, as well as the richer peptide diversity of B. granulifera in relation to S. helianthus. The transcriptomic analysis of B. granulifera, performed by massive cDNA sequencing with 454 pyrosequencing approach allowed the discovery of five new APETx-like peptides (U-AITX-Bg1a-e - including the full sequences of their precursors for four of them), which together with type 1 sea anemone sodium channel toxins constitute a very distinguishable feature of studied sea anemone species belonging to genus Bunodosoma. The molecular modeling of these new APETx-like peptides showed a distribution of positively charged and aromatic residues in putative contact surfaces as observed in other animal toxins. On the other hand, they also showed variable electrostatic potentials, thus suggesting a docking onto their targeted channels in different spatial orientations. Moreover several crab paralyzing toxins (other than U-AITX-Bg1a-e), which induce a variety of symptoms in crabs, were isolated. Some of them presumably belong to new classes of crab-paralyzing peptide toxins, especially those with molecular masses below 2 kDa, which represent the smallest peptide toxins found in sea anemones. (C) 2011 Elsevier Inc. All rights reserved.

Crotamine Pharmacology Revisited: Novel Insights Based on the Inhibition of K-V Channels

Crotamine, a 5-kDa peptide, possesses a unique biological versatility. Not only has its cell-penetrating activity become of clinical interest but, moreover, its potential selective antitumor activity is of great pharmacological importance. In the past, several studies have attempted to elucidate the exact molecular target responsible for the crotamine-induced skeletal muscle spasm. The aim of this study was to investigate whether crotamine affects voltage-gated potassium (K-V) channels in an effort to explain its in vivo effects. Crotamine was studied on ion channel function using the two-electrode voltage clamp technique on 16 cloned ion channels (12 K-V channels and 4 Na-V channels), expressed in Xenopus laevis oocytes. Crotamine selectively inhibits K-V 1.1, K-V 1.2, and K-V 1.3 channels with an IC50 of similar to 300 nM, and the key amino acids responsible for this molecular interaction are suggested. Our results demonstrate for the first time that the symptoms, which are observed in the typical crotamine syndrome, may result from the inhibition of K-V channels. The ability of crotamine to inhibit the potassium current through K-V channels unravels it as the first snake peptide with the unique multifunctionality of cell-penetrating and antitumoral activity combined with K-V channel-inhibiting properties. This new property of crotamine might explain some experimental observations and opens new perspectives on pharmacological uses.

Taxonomy, Hemipenial Morphology, and Natural History of Two Poorly Known Species of Anadia (Gymnophthalmidae) from Northern South America

Anadia pariaensis Rivas, La Marca, and Oliveros, 1999, and Anadia steyeri Nieden, 1914, are two particularly rare and poorly known lizards described from single specimens. In the case of A. pariaensis, it remains known from the holotype, whereas A. steyeri is known from three additional specimens reported in the literature after the original description of the species. A single new specimen of A. pariaensis and five of A. steyeri, including the first adult males recorded for both species, make possible a more representative description of both species, including descriptions of the hemipenes. Despite both species presenting some similar morphological characteristics, the examination of the hemipenial morphology revealed very different organs. The hemipenis of A. steyeri presents some characteristics that resemble the organs of two species from the Santa Marta Mountain Range in the "bitaeniata-group" (Anadia pulchella and Anadia altaserrania). On the other hand, the hemipenes of A. pariaensis are unique morphologically and cannot be associated with the hemipenes known from other species in the genus. We describe variation within both species, and we comment on possible sexual dimorphism (number and arrangement of the femoral pores), natural history, and the known geographic distribution of the species. We also comment on Anadia bumanguesa Rueda-Almonacid and Caicedo 2004 based on a new specimen, the second known. This species may be a synonym of A. steyeri.

A new Fenestrulina (Bryozoa, Cheilostomata) commensal with tube-dwelling anemones (Cnidaria, Ceriantharia) in the tropical southwestern Atlantic

A new species of cheilostome bryozoan, Fenestrulina commensalis n. sp., was collected in December 2008 by scuba at 5–10 meters depth at Guaibura Beach, Guarapari, Espírito Santo state, southeastern Brazil. The specimen was found associated with tubes of the cerianthid Pachycerianthus sp., representing the first commensal association between a bryozoan and a tube-dwelling anemone. Fenestrulina commensalis n. sp. is the third species of the genus found in Brazilian waters; it is distinguished from other Atlantic species of Fenestrulina by its small angular orificial condyles, a single oral spine and basal anchoring rhizoids arising from abfrontal pore chambers. Morphological adaptations to encrust the tubes of cerianthids include anchoring rootlets and weakly contiguous zooids. These morphological features allow the colony the flexibility to grow around the tube and feed relatively undisturbed by silt and detritus, being raised well above the softsediment substratum in which the tube-anemone grows.