A new dwarf species of Proceratophrys Miranda-Ribeiro, 1920 (Anura, Cycloramphidae) from the highlands of Chapada Diamantina, Bahia, Brazil

A new species of Proceratophrys is described from the highlands of northeastern Brazil. Molecular and morphological data suggests that Proceratophrys redacta sp. nov. is sister to P. minuta, and related to P. schirchi and P. cristiceps. The new species is diagnosed by its small size, absence of rostral and palpebral appendages, sagittal ridges interrupted, absence of postocular swellings, snout vertical in profile and dorsal coloration lacking distinct ocelli. The new species represents another example of endemism for the genus Proceratophrys in Chapada Diamantina region, and of another appendageless small-sized species associated with highlands. The phylogenetic results indicate that current morphological groupings in Proceratophrys may not represent natural groups.

Climatic oscillations shape the phylogeographical structure of Atlantic Forest fire-eye antbirds (Aves: Thamnophilidae)

In the present study, mitochondrial (mt)DNA sequence data were used to examine the genetic structure of fire-eye antbirds (genus Pyriglena) along the Atlantic Forest and the predictions derived from the river hypothesis and from a Last Glacial Maximum Pleistocene refuge paleomodel were compared to explain the patterns of genetic variation observed in these populations. A total of 266 individuals from 45 populations were sampled over a latitudinal transect and a number of phylogeographical and population genetics analytical approaches were employed to address these questions. The pattern of mtDNA variation observed in fire-eye antbirds provides little support for the view that populations were isolated by the modern course of major Atlantic Forest rivers. Instead, the data provide stronger support for the predictions of the refuge model. These results add to the mounting evidence that climatic oscillations appear to have played a substantial role in shaping the phylogeographical structure and possibly the diversification of many taxa in this region. However, the results also illustrate the potential for more complex climatic history and historical changes in the geographical distribution of Atlantic Forest than envisioned by the refuge model. (c) 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 105, 900824.

Water balance and spatial distribution of an anuran community from Brazil

Terrestrial amphibians may dehydrate when exposed to low humidity, representing an important factor affecting spatial distribution and community composition. In this study we investigated whether rates of dehydration and rehydration are able to explain the spatial distribution of an anuran community in a Restinga environment at the northern coast of the State of Bahia, Brazil, represented by 11 species distributed in 27 sample units. The environmental data set containing 20 variables was reduced to a few synthetic axes by principal component analysis (PCA). Physiological variables measured were rates of dehydration, rehydration from water, and rehydration from a neutral substrate. Multiple regression analyses were used to test the null hypothesis of no association between the environmental data set (synthetic axes of PCA) and each axis representative of a physiological variable, which was rejected (P < 0.001). Of 15 possible partial regressions only rehydration rate from neutral substrate vs. PC1. and PC2, rehydration rate from water vs. PC1, and dehydration rate vs. PC2 were significant. Our analysis was influenced by a gradient between two different groups of sample units: a beach area with high density of bromeliads and an environment without bodies of water with low density of bromeliads. Species of very specific natural history and morphological characters occur in these environments: Phyllodytes melanomystax and Scinax auratus, species frequently occurring in terrestrial bromeliads, and Ischnocnema paulodutrai, common along the northern coast of Bahia and usually found in forest remnants within environments with low number of bodies of water. In dry environments species with lower rates of dehydration were dominant, whereas species showing greater rates of dehydration were found predominantly in microhabitats with greater moisture or abundance of bodies of water.

Embryonic development of the fossorial gymnophthalmid lizards Nothobachia ablephara and Calyptommatus sinebrachiatus

The evolutionary history of the lizard family Gymnophthalmidae is characterized by several independent events of morphological modifications to a snake-like body plan, such as limb reduction, body elongation, loss of external ear openings, and modifications in skull bones, as adaptive responses to a burrowing and fossorial lifestyle. The origins of such morphological modifications from an ancestral lizard-like condition can be traced back to evolutionary changes in the developmental processes that coordinate the building of the organism. Thus, the characterization of the embryonic development of gymnophthalmid lizards is an essential step because it lays the foundation for future studies aiming to understand the exact nature of these changes and the developmental mechanisms that could have been responsible for the evolution of a serpentiform (snake-like) from a lacertiform (lizard-like) body form. Here we describe the post-ovipositional embryonic development of the fossorial species Nothobachia ablephara and Calyptommatus sinebrachiatus, presenting a detailed staging system for each one, with special focus on the development of the reduced limbs, and comparing their development to that of other lizard species. The data provided by the staging series are essential for future experimental studies addressing the genetic basis of the evolutionary and developmental variation of the Gymnophthalmidae. (C) 2012 Elsevier GmbH. All rights reserved.

Six species of Amazonian Woodcreepers (Aves: Dendrocolaptidae) preying upon lizards and frogs

Dietary data from a large sample of woodcreepers (16 spp., n?=?139), revealed that six species of dendrocolaptids occasionally feed upon lizards and frogs. These birds, which are mainly insectivorous, encounter and feed on lizards while perching on tree trunks, probably in association with army-ant swarm feeding behaviour. Frog intake may be related to declines in the abundance of invertebrate prey. The bones recovered were identified as one small species of gecko, Gonatodes humeralis, and at least one anuran. We estimate that in the entire sample, about eight lizards and two frogs were ingested. The partially digested gecko material allows determination of which bones are more resistant to digestion, although it is possible that these elements were differentially retained in the stomach. These elements correspond to the more frequently preserved bones in the fossil record of geckos, indicating that the same portions of the skeleton persist under the processes of both digestion and fossilization.

Effects of intravenous furosemide on mucociliary transport and rheological properties of patients under mechanical ventilation

The use of intravenous (IV) furosemide is common practice in patients under mechanical ventilation (MV), but its effects on respiratory mucus are largely unknown. Furosemide can affect respiratory mucus either directly through inhibition of the NaK(Cl)2 co-transporter on the basolateral surface of airway epithelium or indirectly through increased diuresis and dehydration. We investigated the physical properties and transportability of respiratory mucus obtained from 26 patients under MV distributed in two groups, furosemide (n = 12) and control (n = 14). Mucus collection was done at 0, 1, 2, 3 and 4 hours. The rheological properties of mucus were studied with a microrheometer, and in vitro mucociliary transport (MCT) (frog palate), contact angle (CA) and cough clearance (CC) (simulated cough machine) were measured. After the administration of furosemide, MCT decreased by 17 ± 19%, 24 ± 11%, 18 ± 16% and 18 ± 13% at 1, 2, 3 and 4 hours respectively, P < 0.001 compared with control. In contrast, no significant changes were observed in the control group. The remaining parameters did not change significantly in either group. Our results support the hypothesis that IV furosemide might acutely impair MCT in patients under MV.

Screening bacterial metabolites for inhibitory effects against Batrachochytrium dendrobatidis using a spectrophotometric assay

Certain bacteria present on frog skin can prevent infection by the pathogenic fungus Batrachochytrium dendrobatidis (Bd), conferring disease resistance. Previous studies have used agar-based in vitro challenge assays to screen bacteria for Bd-inhibitory activity and to identify candidates for bacterial supplementation trials. However, agar-based assays can be difficult to set up and to replicate reliably. To overcome these difficulties, we developed a semi-quantitative spectrophotometric challenge assay technique. Cell-free supernatants were prepared from filtered bacterial cultures and added to 96-well plates in replicated wells containing Bd zoospores suspended in tryptone-gelatin hydrolysate-lactose (TGhL) broth medium. Plates were then read daily on a spectrophotometer until positive controls reached maximum growth in order to determine growth curves for Bd. We tested the technique by screening skin bacteria from the Australian green-eyed tree frog Litoria serrata. Of bacteria tested, 31% showed some degree of Bd inhibition, while some may have promoted Bd growth, a previously unknown effect. Our cell-free supernatant challenge assay technique is an effective in vitro method for screening bacterial isolates for strong Bd-inhibitory activity. It contributes to the expanding field of bioaugmentation research, which could play a significant role in mitigating the effects of chytridiomycosis on amphibians around the world.

Amphibian biodiversity in Taita Hills, Kenya

Species richness varies greatly across geographical regions. Eastern Arc Mountains (EAM) of Kenya and Tanzania is one of the global biodiversity hotspots. Despite this, high species diversity the explanatory factors have remained largely unexplored. Herein, this study first investigated amphibian species richness patterns in the EAM and particularly the reasons for the low richness in Taita Hills. It tested the hypothesis that the low richness is due to past forest loss or other factors. The results demonstrated that the regional species richness pattern was influenced largely by mean annual rainfall and not forest area. Secondly, using the 26 currently recorded amphibians in the Taita Hills, it investigated the relationship between amphibian species composition along anthropogenic habitat disturbance and elevation gradients. It tested the hypothesis that sites with similar environmental characteristics (temperature, rainfall and elevation), in close proximity and with similar disturbance levels (habitat types) harbour similar species composition. It was found that amphibian species composition differed in terms of elevation and was explained by both temperature and rainfall. Therefore sites with similar environmental characteristics, disturbance levels and in close proximity geographically have similar amphibian composition. Thirdly, diagnostic characters, distribution, basic life history characteristics and conservation status of all currently known amphibians in the Taita Hills were provided. Finally, first long term life history and ecological characteristics of a brevicipitid frog (Callulina sp) was provided. The results showed that this frog abundance and distribution is influenced mainly by mean monthly temperature, breeds during the long dry season and exhibit parental care. Results of this study strongly recommend increasing indigenous forest cover in order to enhance the conservation of the endemic indigenous forest associated amphibians such as Callulina sp, Boulengerula taitana and Boulengerula niedeni.

Identification and genetic diversity in phytoplasmas associated with diseases of cassava and other agronomic relevant crops in south-east Asia and Latin America

Identification and genetic diversity of phytoplasmas infecting tropical plant species, selected among those most agronomically relevant in South-east Asia and Latin America were studied. Correlation between evolutionary divergence of relevant phytoplasma strains and their geographic distribution by comparison on homologous genes of phytoplasma strains detected in the same or related plant species in other geographical areas worldwide was achieved. Molecular diversity was studied on genes coding ribosomal proteins, groEL, tuf and amp besides phytoplasma 16S rRNA. Selected samples infected by phytoplasmas belonging to diverse ribosomal groups were also studied by in silico RFLP followed by phylogenetic analyses. Moreover a partial genome annotation of a ‘Ca. P. brasiliense’ strain was done towards future application for epidemiological studies. Phytoplasma presence in cassava showing frog skin (CFSD) and witches’ broom (CWB) diseases in Costa Rica - Paraguay and in Vietnam – Thailand, respectively, was evaluated. In both cases, the diseases were associated with phytoplasmas related to aster yellows, apple proliferation and “stolbur” groups, while only phytoplasma related to X-disease group in CFSD, and to hibiscus witches’ broom, elm yellows and clover proliferation groups in CWB. Variability was found among strains belonging to the same ribosomal group but having different geographic origin and associated with different disease. Additionally, a dodder transmission assay to elucidate the role of phytoplasmas in CWB disease was carried out, and resulted in typical phytoplasma symptoms in periwinkle plants associated with the presence of aster yellows-related strains. Lethal wilt disease, a severe disease of oil palm in Colombia that is spreading throughout South America was also studied. Phytoplasmas were detected in symptomatic oil palm and identified as ‘Ca. P. asteris’, ribosomal subgroup 16SrI-B, and were distinguished from other aster yellows phytoplasmas used as reference strains; in particular, from an aster yellows strain infecting corn in the same country.

Die Schleimkeratine TK-alpha and TK-gamma des Zebrabärblings Danio rerio

Die Schleimkeratine TKα und TKγ aus dem Schleimaal Eptatretus stoutii besitzten für Keratine außergewöhnliche Eigenschaften. In speziellen Drüsen reifen die Schleimkeratine zu 3 µm dicken und bis zu 60 cm langen kabelartigen Filamenten heran und werden anschließend zur Feindabwehr ins umgebende Wasser extrazellulär sezerniert, wodurch die viskoelastischen Eigenschaften des Schleims modifiziert werden. Mittlerweile wurden die Schleimkeratine auch in höheren Wirbeltiergruppen (Knochenfische und Amphibien) entdeckt. Zu Beginn meiner Promotion war jedoch bis auf EST-Verteilungsprofile noch nichts über die Expression und Funktion der Schleimkeratine in diesen Organismen bekannt. rnIm Rahmen meiner Arbeit wurden die Schleimkeratine TKα und TKγ erstmalig im Zebrabärbling Danio rerio identifiziert und näher charakterisiert. Mittels rekombinanter Expression wurden TKα und TKγ in ausreichenden Mengen hergestellt und auf ihre Bindungseigenschaften hin untersucht. Hierbei konnte ich zeigen, dass TKα und TKγ einerseits miteinander Heteromere formen und andererseits, dass das TKα in der Lage ist, auch homopolymere Strukturen auszubilden. Letztere Eigenschaft wurde bisher noch bei keinem bekannten cytoplasmatischen Keratin beschrieben. Ergänzend zu diesen Untersuchungen wurde eine Expressionsanalyse durchgeführt. Hierbei konnte gezeigt werden, dass die Schleimkeratine im Zebrabärbling nicht extrazellulär sezerniert werden und zum anderen keine höheren, kabelartigen Strukturen ausformen. Vielmehr werden die Schleimkeratine bei adulten Tieren in den basalen Zellschichten der Epidermis exprimiert, welche keinen mechanischen Schutz in Form von Schuppen aufweisen (Stirnhautepidermis, Epidermis in Geweben zwischen den Flossenstrahlen). Innerhalb dieser Zellen formen die Schleimkeratine ein filamentöses Netzwerk aus, dass sich an der basalen Zellseite konzentriert. Eine mögliche Funktion von TKα und TKγ könnte demnach in der Erhöhung der mechanischen Integrität von stark beanspruchten Geweben liegen, die keinen Schutz in Form von Schuppen aufweisen. So werden TKα und TKγ in larvalen Entwicklungsstadien in der Epidermis, sowie im mechanisch stark beanspruchten Notochord koexprimiert. rnDa das Notochord im Zebrabärbling auch in entwicklungsbiologischen Vorgängen eine entscheidende Rolle spielt und weiterhin in aktuellen Untersuchungen am glatten Krallen-frosch Xenopus laevis Funktionen der Schleimkeratine TKα und TKγ innerhalb von Degenerationsprozessen während der Metamorphose nachgewiesen werden konnten, sind auch im Zebrabärbling Danio rerio Funktionen der Schleimkeratine TKα und TKγ im Rahmen von Entwicklungsprozessen denkbar.rn

Wet bioadhesion in tree frogs

Viele Tiere wie etwa Geckos oder Laubfrösche können mittels ihrer Haftscheiben an Oberflächen kleben. Diese Haftscheiben ermöglichen es den Tieren, sich während ihrerrnFortbewegung an Oberflächen anzuheften und wieder zu lösen unabhängig von denrnvorherrschenden Umweltbedingungen. Frösche besitzen mikro- und nanostrukturierternsowie charakteristisch geformte Haftscheiben an Finger- und Zehenenden. Ihre besonderernevolutionäre Errungenschaft, sich stark und zugleich reversibel in sowohl trockenen alsrnauch feuchten Umgebungen anzuhaften, hat die Wissenschaft zur Nachahmung und Untersuchungrndieser Strukturen inspiriert. Zum besseren Verständnis der Mechanismen vonrnAnhaftung und Loslösung bei Laubfröschen wurden weiche, elastische und mikrostrukturierternOberflächen hergestellt, indem PDMS (Polydimethylsiloxan) auf einer Siliziummaskernmit Hexagonstruktur aufgetragen und vernetzt wurde. Dadurch wurden Anordnungenrnvon hexagonalen Mikrosäulen mit spezifischen geometrischen Eigenschaften undrnunterschiedlichen Kontaktgeometrien (normale, flache Form, T-Form und konkave Formrnder Säulenenden) erhalten. Um den Einfluss der van-der-Waals, hydrodynamischen,rnKapillar-und Adhäsionskräfte zu verstehen, wurden verschiedene experimentelle Ansätzernverfolgt: Die auf eine einzelne Säule wirkenden Adhäsionskräfte wurden mittelsrnRasterkraftmikroskopie gemessen. Dazu wurden speziell hergestellte kolloidale Sensorenrnverwendet. Diese Experimente wurden sowohl mit als auch ohne Flüssigkeitsfilm auf derrnSäule durchgeführt. Die Ergebnisse zeigten den Beitrag von Kapillarkraft und direktenrnKontaktkräften zur Adhäsionskraft bei Vorliegen eines Flüssigkeitsfilms. Die Adhäsionrnfiel umso größer aus, je weniger Flüssigkeit zwischen Sensor und Säule vorhanden war.rnIm Falle einer trockenen Adhäsion zeigte die Säule mit T-Form die höchste Adhäsion. Darndie Haftscheiben der Laubfrösche weich sind, können sie dynamisch ihre Form ändern,rnwas zu einer Änderung der hydrodynamischen Kraft zwischen Scheibe und Oberflächernführt. Der Einfluss der Oberflächenverformbarkeit auf die hydrodynamische Kraft wurderndaher am Modellsystem einer Kugel untersucht, welche sich einer weichen und ebenenrnOberfläche annähert. Dieses System wurde sowohl theoretisch über die Simulation finiterrnElemente als auch experimentell über die Messung mit kolloidalen Sonden untersucht.rnSowohl experimentelle Ergebnisse als auch die Simulationen ergaben eine Abnahme derrnhydrodynamischen Kraft bei Annäherung des kolloidalen Sensors an eine weiche undrnelastische Oberfläche. Beim Entfernen der Sensors von der Oberfläche verstärkte sichrndie hydrodynamische Anziehungskraft. Die Kraft, die zur Trennung eines Partikels von einer Oberfläche in Flüssigkeit notwendig ist, ist für weiche und elastischen Oberflächenrngrößer als für harte Oberflächen. In Bezug zur Bioadhäsion bei Laubfröschen konnternfestgestellt somit festgestellt werden, dass sich der hydrodynamische Anteil zur feuchtenrnBioadhäsion aufgrund der weichen Oberfläche erhöht. Weiterhin wurde der Einflussrndes Aspektverhältnisses der Säulen auf die Reibungskraft mittels eines kolloidalen Sensorsrnuntersucht. Gestreckte Säulen zeigten dabei eine höhere Reibung im Vergleich zu.rnSäulen mit einem gestreckten Hexagon als Querschnitt.

Populationsgenetik der ersten Bauern Mitteleuropas

Die vorliegende Dissertation ist eine molekulargenetische Studie an humanem neolithischem Skelettmaterial. Im zentralen Blickpunkt stand die Bestimmung der Variabilität der mitochondrialen Haplogruppen einer frühneolithischen Stichprobe aus drei unterschiedlichen Kulturkreisen, welche die Linearbandkeramik (LBK und AVK), die Körös-Kultur und eine Sammelkategorie osteuropäischer spätmeso- und frühneolithischer Kulturen umfasste. Im Vergleich dieser Gruppen untereinander sowie mit Rezentdaten moderner Populationen aus vergleichbaren Gebieten Mittel- und Osteuropas sowie dem Nahen Osten sollten bestehende Modelle und Hypothesen zur Neolithisierung Mitteleuropas geprüft werden. Insgesamt konnte für 43 neolithische Individuen aus 16 Fundorten der reproduzierbare Nachweis endogener DNA erbracht werden. Eine eindeutige Haplogruppenbestimmung konnte durch die Sequenzierung vier überlappender Fragmente der mitochondrialen Hypervariablen Region I sowie durch RFLP-Analyse zusätzlicher charakteristischer Nukleotidpositionen für alle 43 Individuen durchgeführt werden. Die neolithischen Individuen der Linearbandkeramik sowie der Körös-Kultur zeigten eine hohe Diversität an bekannten europäischen Haplogruppen, wohingegen die kleinere Stichprobe aus dem Gebiet Osteuropas eine auffällige Homogenität aufwies. Neben Frequenzunterschieden zur modernen mitteleuropäischen Bevölkerung war innerhalb der LBK/AVK-Stichprobe eine hohe Frequenz der Haplogruppe N1a festzustellen, welche nicht in den beiden anderen neolithischen Stichproben zu finden war und auch in der heutigen Rezentbevölkerung Eurasiens und Nordafrikas nur mit einer durchschnittlichen Frequenz von 0,2% vertreten ist. Innerhalb der Individuen der Körös-Kultur fanden sich zwei Haplotypen, welche heute nicht in Europa bekannt sind, dagegen jedoch in Süd- bzw. Nordostasien gehäuft vorkommen. Die Ergebnisse der aDNA-Analysen bestätigten im Wesentlichen das komplexe Bild der Neolithischen Transition in Mitteleuropa und konnten die, für diesen Raum postulierte, Hypothese der leap frog colonization weitestgehend unterstützen. Auch für den geographischen Vergleichsraum des nördlichen Osteuropa konnten Parallelen zur etablierten Sichtweise der archäologischen Forschung zu diesem Gebiet und den vorliegenden Ergebnissen der aDNA-Analysen aufgezeigt werden. Die zeitlich verzögerte Annahme der neolithischen Lebensweise im waldreichen nördlichen Osteuropa spiegelt sich in der reduzierten Diversität an mtDNA-Haplogruppen wider. Die vorliegende Dissertation konnte nicht nur durch die Ergebnisse der Haplogruppen-Bestimmung, sondern vor allem durch die umfangreichen und elaborierten Reproduktions- und Authentifizierungprotokolle deutlich machen, dass der Nachweis von humaner alter DNA trotz der allgegenwärtigen, methodenimmanenten Kontaminationsgefahr unter streng kontrollierten Bedingungen möglich ist. Gleichermaßen konnte veranschaulicht werden, dass die aDNA-Analyse wertvolle Hinweise auf das genetische status quo einer Population liefern kann, welche nicht bzw. nur in sehr eingeschränkten Maße von rezenten DNA-Daten abgeleitet werden können. Als sekundäres Ergebnis erlaubte der bislang größte vorliegende Datensatz von ~2500 Klonsequenzen zudem einen detaillierten Einblick in Häufigkeiten und Verteilungsmuster von post mortem Sequenzveränderungen. Es konnten für den mitochondrialen Bereich der Nukleotidpositionen 15997-16409 so genannte hot bzw. cold spots definiert werden, welche für die Auswertung und Interpretation von zukünftigen Sequenzierungen der Hypervariablen Region I des mt-Genoms von entscheidender Bedeutung sein werden.

[(99m)Tc]Demomedin C, a radioligand based on human gastrin releasing peptide(18-27): synthesis and preclinical evaluation in gastrin releasing peptide receptor-expressing models

The synthesis and preclinical evaluation of [(99m)Tc]Demomedin C in GRPR-expressing models are reported. Demomedin C resulted by coupling a Boc-protected N(4)-chelator to neuromedin C (human GRP(18-27)), which, after (99m)Tc-labeling, afforded [(99m)Tc]Demomedin C. Demomedin C showed high affinity and selectivity for the GRPR during receptor autoradiography on human cancer samples (IC(50) in nM: GRPR, 1.4 ± 0.2; NMBR, 106 ± 18; and BB(3)R, >1000). It triggered GRPR internalization in HEK-GRPR cells and Ca(2+) release in PC-3 cells (EC(50) = 1.3 nM). [(99m)Tc]Demomedin C rapidly and specifically internalized at 37 °C in PC-3 cells and was stable in mouse plasma. [(99m)Tc]Demomedin C efficiently and specifically localized in human PC-3 implants in mice (9.84 ± 0.81%ID/g at 1 h pi; 6.36 ± 0.85%ID/g at 4 h pi, and 0.41 ± 0.07%ID/g at 4 h pi block). Thus, human GRP-based radioligands, such as [(99m)Tc]Demomedin C, can successfully target GRPR-expressing human tumors in vivo while displaying attractive biological features--e.g. higher GRPR-selectivity--vs their frog-homologues.

A venom-derived neurotoxin, CsTx-1, from the spider Cupiennius salei exhibits cytolytic activities

CsTx-1, the main neurotoxic acting peptide in the venom of the spider Cupiennius salei, is composed of 74 amino acid residues, exhibits an inhibitory cysteine knot motif, and is further characterized by its highly cationic charged C terminus. Venom gland cDNA library analysis predicted a prepropeptide structure for CsTx-1 precursor. In the presence of trifluoroethanol, CsTx-1 and the long C-terminal part alone (CT1-long; Gly-45-Lys-74) exhibit an α-helical structure, as determined by CD measurements. CsTx-1 and CT1-long are insecticidal toward Drosophila flies and destroys Escherichia coli SBS 363 cells. CsTx-1 causes a stable and irreversible depolarization of insect larvae muscle cells and frog neuromuscular preparations, which seem to be receptor-independent. Furthermore, this membranolytic activity could be measured for Xenopus oocytes, in which CsTx-1 and CT1-long increase ion permeability non-specifically. These results support our assumption that the membranolytic activities of CsTx-1 are caused by its C-terminal tail, CT1-long. Together, CsTx-1 exhibits two different functions; as a neurotoxin it inhibits L-type Ca(2+) channels, and as a membranolytic peptide it destroys a variety of prokaryotic and eukaryotic cell membranes. Such a dualism is discussed as an important new mechanism for the evolution of spider venomous peptides.

Meprins, membrane-bound and secreted astacin metalloproteinases

The astacins are a subfamily of the metzincin superfamily of metalloproteinases. The first to be characterized was the crayfish enzyme astacin. To date more than 200 members of this family have been identified in species ranging from bacteria to humans. Astacins are involved in developmental morphogenesis, matrix assembly, tissue differentiation and digestion. Family members include the procollagen C-proteinase (BMP1, bone morphogenetic protein 1), tolloid and mammalian tolloid-like, HMP (Hydra vulgaris metalloproteinase), sea urchin BP10 (blastula protein) and SPAN (Strongylocentrotus purpuratus astacin), the 'hatching' subfamily comprising alveolin, ovastacin, LCE, HCE ('low' and 'high' choriolytic enzymes), nephrosin (from carp head kidney), UVS.2 from frog, and the meprins. In the human and mouse genomes, there are six astacin family genes (two meprins, three BMP1/tolloid-like, one ovastacin), but in Caenorhabditis elegans there are 40. Meprins are the only astacin proteinases that function on the membrane and extracellularly by virtue of the fact that they can be membrane-bound or secreted. They are unique in their domain structure and covalent subunit dimerization, oligomerization propensities, and expression patterns. They are normally highly regulated at the transcriptional and post-translational levels, localize to specific membranes or extracellular spaces, and can hydrolyse biologically active peptides, cytokines, extracellular matrix (ECM) proteins and cell-surface proteins. The in vivo substrates of meprins are unknown, but the abundant expression of these proteinases in the epithelial cells of the intestine, kidney and skin provide clues to their functions.