A NOVEL PLASMINOGEN-ACTIVATOR FROM SNAKE-VENOM - PURIFICATION, CHARACTERIZATION, AND MOLECULAR-CLONING

A novel plasminogen activator from Trimeresurus stejnegeri venom (TSV-PA) has been identified and purified to homogeneity. It is a single chain glycoprotein with an apparent molecular weight of 33,000 and an isoelectric point of pH 5.2. It specifically activates plasminogen through an enzymatic reaction. The activation of human native GIu-plasminogen by TSV-PA is due to a single cleavage of the molecule at the peptide bond Arg(561)-Val-(562). Purified TSV-PA, which catalyzes the hydrolysis of several tripeptide p-nitroanilide substrates, does not activate nor degrade prothrombin, factor X, or protein C and does not clot fibrinogen nor show fibrino(geno)lytic activity in the absence of plasminogen. The activity of TSV-PA was readily inhibited by phenylmethanesulfonyl fluoride and by p-nitrophenyl-p-guanidinobenzoate. Oligonucleotide primers designed on the basis of the N-terminal and the internal peptide sequences of TSV-PA were used for the amplification of cDNA fragments by polymerase chain reaction. This allowed the cloning of a full-length cDNA encoding TSV-PA from a cDNA library prepared from the venom glands. The deduced complete amino acid sequence of TSV-PA indicates that the mature TSV-PA protein is composed of 234 amino acids and contains a single potential N-gIycosylation site at Asn(1G1). The sequence of TSV-PA exhibits a high degree of sequence identity with other snake venom proteases: 66% with the protein C activator from Aghistrodon contortrix contortrix venom, 63% with batroxobin, and 60% with the factor V activator from Russell's viper venom. On the other hand, TSV-PA shows only 21-23% sequence similarity with the catalytic domains of u-PA and t-PA. Furthermore, TSV-PA lacks the sequence site that has been demonstrated to be responsible for the interaction of t-PA (KHRR) and u-PA (RRHR) with plasminogen activator inhibitor type 1.

A horsefly saliva antigen 5-like protein containing RTS motif is an angiogenesis inhibitor

The Ag5 proteins are the most abundant and immunogenic proteins in the venom secretory ducts of stinging insects. An antigen 5-like protein (named tabRTS) composed of 221 amino acid residues was purified and characterized from the salivary glands of the horsefly, Tabanus yao (Diptera, Tabanidae). Its cDNA was cloned from the cDNA library of the horsefly's salivary gland. TabRTS containing the SCP domain (Sc7 family of extracellular protein domain) was found in insect antigen 5 proteins. More interestingly, there is an Arg-Thr-Ser (RTS) disintegrin motif at the C-terminus of tabRTS. The RTS motif is positioned in a loop bracketed by cysteine residues as those found in RTS-disintegrins of Crotalidae and Viperidae snake venoms, which act as angiogenesis inhibitors. Endothelial Cell Tube formation assay in vitro and chicken chorioallantoic membrane (CAM) angiogenesis assay in vivo were performed as to investigate the effect of tabRTS on angiogenesis. It was found that tabRTS could significantly inhibit angiogenesis in vitro and in vivo. Anti-alpha(1)beta(1) monoclonal antibody could dose-dependently inhibit the anti-angiogenic activity of tabRTS. This result indicated that tabRTS possibly targets the alpha(1)beta(1) integrin to exert the anti-angiogenic activity as snake venom RTS-/KTS-disintegrins do. The current work revealed the first angiogenesis inhibitor protein containing RTS motif from invertebrates, a possible novel type of RTS-disintegrin. (C) 2009 Elsevier Ltd. All rights reserved.

A unique group of inactive serine protease homologues from snake venom

A number of inactive serine protease homologues (SPHs), which have poorly understood functions, have been identified in invertebrates and vertebrates. Recently, several SPH transcripts have been reported from snake venom glands, which provide potential ne

A novel serine protease inhibitor from Bungarus fasciatus venom

By Sephadex G-50 gel filtration, cation-exchange CM-Sephadex C-25 chromatography and reversed phase high-performance liquid chromatography (HPLC), a novel serine protease inhibitor named bungaruskunin was purified and characterized from venom of Bungarus fasciatus. Its cDNA was also cloned from the cDNA library of B. fasciatus venomous glands. The predicted precursor is composed of 83 amino acid (aa) residues including a 24-aa signal peptide and a 59-aa mature bungaruskunin. Bungaruskunin showed maximal similarity (64%) with the predicted serine protease inhibitor blackelin deduced from the cDNA sequence of the red-bellied black snake Pseudechis porphyriacus. Bungaruskunin is a Kunitz protease inhibitor with a conserved Kunitz domain and could exert inhibitory activity against trypsin, chymotrypsin, and elastase. By screening the cDNA library, two new B chains of beta-bungarotoxin are also identified. The overall structures of bungaruskunin and beta -bungarotoxin B chains are similar; especially they have highly conserved signal peptide sequences. These findings strongly suggest that snake Kunitz/BPTI protease inhibitors and neurotoxic homologs may have originated from a common ancestor. (c) 2007 Elsevier Inc. All rights reserved.

Indian squid, Uroteuthis duvauceli, is an important commercial marine fauna in the coastal waters of Oman Sea

About 3600 specimens were collected by bottom trawl at 15 sampling stations. 24 biometric characters were measured for each specimens at the laboratory.. Microscopic cross – sections of statolith were used for age determination. Sex determination and fecundity were determined. Population dynamics parameters as well as stock as stock assessment including cohort analysis were estimated using FISAT software. The findings showed that Dorsal Mantle Length (DML) and Body weight (BW) of the Indian squid were 133.9 ± 0.78 mm and 99.61 ± 0.95 g respectively. Strong correlation was found between these 2 variables (R2 = 0.90). The maximum age was 5 years. Relationship between DML and age was highly significantly of p ≤ 0.05. Overall sex ratio (M: F = 0.52) was significantly different from the expected 1:1 ratio (p ≤ 0.05). The ovary weight and nidamental glands weight were 7.72 ± 0.0006 g and 3.07 ± 0.0003g respectively. Absolute and relative fecundity of the Indian squid were found to be 122733 ± 30.87 and 2348 ± 0.4 respectively. GSI were 14.35 in April and 8.63 in July. This squid is therefore a spring spawner. The infinite dorsal mantle length were 258.62 mm for females, 194.72 mm for males and 252.02 for both sexes respectively. For population growth and mortality parameters; K (0.65 per year for both sexes, 0.85 per year for males, 0.65 per year for females); t0 (0.24year for both sexes, 0.22 year in females, 0.26 year in male); φ` (2.30 in both sexes, 2.47 for males, 2.37 for females); Z (1.17 per year for both sexes, 1.10 per year in females, 1.39 per year, in males); M (0.70 per year for both sexes, 0.90 for males, 0.67 for females); F(0.27 per year for both sexes, 0.27 per year in males, 0.195 per year in females). Exploitation coefficient were 0.51 per year for both sexes, 0.57 per year males and 0.51 per year females respectively. The results indicates that since the Indian squid is a short live aquatic organism, therefore, the exploitation coefficient could be raised to 0.7 per year. The analysis showed that total biomass and MSY were 10103.5 ton and 2576.4 ton respectively. These findings are the first study of its sort about the Indian squid in the coastal waters of Oman Sea as well as North-West of Indian Ocean.

Contribución al estudio biológico de Loligo brasiliensis Bl

A total of 592 individuals of Loligo brasiliensis from the Mar del Plata coastal fishing area (Buenos Aires prov., Argentina) have been studied during the 1961-1964 period. From a morphological point of view the population appears to be uniform and homogeneus. A brief description of this species is given in this paper since references in the literature are scarce from the time at which Blainville (1923) first described it. The only further references are found in D'Orbigny (1835), and Ferrusac (1839), and in Hoyle (1886), and Tyron Pilsbry (1879). In this paper the species was mentioned only as a bibliographical reference on morphological or biological conditions has been found in the literature. The distribution of this species ranges from Cuba, Brazil, Uruguay to the Argentine coast, probably down to the Gulf of San Jorge. The samples had been studied with respect to various body measurements by classifing the individuals in total length classes, since body length was considered the most significant measurement. The condition factor K has been calculated for different sexes and ages, for the various length classes. The results lead to the conclusion that the smaller the length the higher is the value obtained for K and viceversa. This is due to the fact that the length of the tentacle increases considerably with increasing size. Since the tentacle are quite light the factor K diminishes accordingly. The condition factor increases considerably from December to April with an average of 0.42, decrease and becomes stable from March to October, with an average of 0.30. This is a consequence of the ripening of the sex glands. The sex-ratios are as following: year 1961, 42 % female, 42 % male; year 1962, 51 % female, 45 % male; year 1963, 46 % female, 53 % male; year 1964, 26 % female, 42 % male, 32 % indif. The great percentage of 72 undifferentiated young individuals in the 1964 (March) sampling increases the ratio of undifferentiation. A short morphological description of both ovules and spermatozoos is given. An examination of the sex glands leads to the following conclusions: a) male and female sex gland in a preparatory stage during the whole year; b) the highest percentage of ripe glands is found through, November-March; e) the spawning appears to precede rather slowly, but this certain since the spawning environment does not coincide with the natural habitat of the species. Few spawning individuals were found; d) sexual differentiation begins at body lenght from 30 to 40 mm; i.e. a total length of approximately 145 mm. At a body length of 70 mm. the hectocotilication (sexual character) begins to appear. In June 1962, a sample gathered at Rawson (Chubut) was analyzed. The conclusion was reached that the sex glands in this population are in an earlier stage of development in comparison with those from the Mar del Plata area. Also the average for the factor K which were found to be 0.17 for females and 0.19 for male, are rather low for that date. These physiological facts are possibly related to morphological differences which will be pointed out in a forthcoming publication. Some very typical associations with Artemesia longinaris and Percophis brasiliensis were found. Cannibalism has been observed.

Taxonomía y distribución de los poliquetos pelágicos del Atlántico Sudoccidental

This work is based on the analysis of 420 planktonic samples of 7 oceanopraphic cruises distributed over the Argentine, Uruguayan and South brasilian continental shelf (SW Atlantic ocean), as well as from some oceanic sectors, adjacent to the continental slope. Vertical hauls were performed in all stations from 100 m depth to surface, except in the Walter Herwig cruise (where vertical hauls were predominantly performed out of slope sectors, between 300 and 500 m depth to surface) and Productividad cruise in which only surface waters were hauled. A list of 27 species are determined, corresponding to 5 families: Iospilidae (3 species), Lopadorrhynchidae (4), Alciopidae (9), Typhloscolecidae (5) and Tomopteridae (6). Larvae and epitokous forms of benthonic species are not taken into account. The genus Iospilus is revised, Pariospilus and Iospilopsis being considered their synonyms; the identity of Pariospilus affinis Viguier is maintained, being transferred to the genus Iospilus. The species Vanadis studeri Apstein is redescribed and its synonymy is established. The taxonomic value of the apical glands of Tomopteris species is discussed and some specimens are found to coincide with T. kefersteini in relation to the mentioned glands. All the species found in this work are described and illustrated, a systematic key being added for their identification. Considering the vertical nature of the hauls, it was not possible to specify the habitats of the different species; for this reason they are grouped as species from subtropical and subantartic areas of influence. The first group, made up of 17 species, shows and evident graduation in its latitudinal distribution, some of them being more restricted in their distribution than the others. The second group, of 4 species, is found south to the tropical convergence, in transitional waters, towards cold sectors. The third group, of 6 species, is found to be distributed all along the continental shelf, in subtropical and subantartic regions, and extending their distribution northwards, possibly related to deep water levels. The general scheme is coincident with the distribution of other planktonic groups (Copepods, Euphausiids). As a general feature, neither coastal nor shelf water specimens of pelagic Polychaeta were found, with exception of T. septentrionalis. A comparison with the results in Tebble's paper (1960) in the southwest Atlantic ocean is made, 12 of our species being coincidently found in the same hydrological area by that author. The drift of the main water masses of the South Atlantic ocean is accepted as a possible cause for the distribution of the pelagic Polychaeta of the southwest Atlantic regions.

Pituitary adenylate cyclase-activating polypeptide

Pituitary adenylate cyclase-activating polypeptide (PACAP) which belongs to the secretin/glucagon/ VIP family has been originally isolated from the sheep hypothalamus on the basis of its ability to stimulate cAMP formation in culture rat anterior pituitary cells. Post-translational processing of the PACAP precursor generates two biologically active molecular forms, PACAP-38 and PACAP-27. The primary structure of PACAP has been remarkably conserved during evolution. The sequence of PACAP-27 exhibits substantial similarities with those of vasoactive intestinal polypeptide (VIP), glucagon and secretin. The gene encoding the PACAP precursor is widely expressed in brain and various peripheral organs, notably in endocrine glands, gastro-intestinal, urogenital tracts and respiratory system. In vivo, and in vitro studies have shown that PACAP exhibits multiple activities especially a trophic activity during ontogenesis, notably in the adrenal medulla and the central nervous system. The biological effects of PACAP are mediated through three distinct receptor subtypes which exhibit differential affinities for PACAP and VIP. The PAC1 receptor, which shows high selectivity for PACAP, is coupled to several transduction systems. In contrast, VPAC1 and VPAC2, which bind with the same affinity for PACAP and VIP, are mainly coupled to the adenylyl cyclase pathway. In conclusion, PACAP is neuropeptide, and it functions as a hypothalamic hormone, neurohormone, neuromodulator, vasodilator, neurotransmitter or trophic factor in the brain and the various organs.

Circulating immunoglobulin A- and immunoglobulin G-secreting hybridoma cells in peripheral blood preferably migrate to female genital tracts. The role of sex hormones

Antigen-specific circulating immunoglobulin-secreting cells (ISC) migrate to various secondary and tertiary lymphoid tissues. To understand the migration of the cells into the genital tract and its regulation by sex hormones, spleen-derived SG2 hybridoma cells secreting immunoglobulin G2b (IgG2b) and Peyer's patch-derived PA4 hybridoma cells secreting polymer IgA were labelled with (3) H-TdR, and intravenously injected into syngeneic mice of both sexes. Using flow cytometry, surface molecular markers of plasma cells, CD38 and CD138, and adhesion molecules, CD49d, CD162, and CD11a were found to be positive in SG2 and PA4 cells, but CD62L, alpha4beta7 and CD44 were not expressed on these cells. The relative distribution indexes (RDIs) of the cells in genital tract and other tissues were measured. The means of RDIs of SG2 and PA4 cells in female genital tissues were 6.5 and 4.5 times as many as the means in male genital tissues, respectively. The treatment of ovariectomized mice with beta-oestradiol significantly increased the RDIs of PA4 cells in cervix and vagina, but decreased the RDIs of SG2 cells in vagina, horn of uterus, uterus and rectum (P <0.05). Progesterone treatment increased the RDIs of PA4 cells in vagina and rectum (P <0.05). The treatment with testosterone significantly increased the RDIs of SG2 and PA4 cells in epididymis and accessory sex glands (P <0.05). These results demonstrate that the female genital tract is the preferable site for the migration of circulating hybridoma cells to the male genital tract, and sex hormones play an important role in regulation of the migration of circulating ISC to genital tracts.

Fridericia liangi sp nov (Enchytraeidae, Oligochaeta) from Changbaishan Mountain, China

We describe one new enchytraeid species, Fridericia liangi sp. nov., from Mt. Changbaishan, Jilin Province, northeastern China. It was collected from soils at the foot of Changbaishan Mountain and is distinguished from all known congeners by the following combination of characters: 1) no lateral chaetae, only ventral chaetae throughout, 2) a maximum of four chaetae in ventral preclitellar bundles, 3) one chaeta in ventral postclitellar bundles, 4) dorsal pores from VII on, 5) esophageal appendages unbranched, 6) coelomocytes without refractile vesicles, 7) clitellum girdle shaped, well developed, 8) no subneural glands, and 9) spermathecae simple.

Three new species of Tubificinae, Oligochaeta, from two plateau lakes in Southwest China

Three new species of Tubificinae (Naididae, Oligochaeta), Varichaetadrilus vestibulatus n. sp., Aulodrilus apeniatus n. sp., and Ilyodrilus mesoprostatus n. sp., are reported from Fuxian Lake and Xingyun Lake of Yunnan Province, Southwest China. V. vestibulatus differs from its allies by possessing modified spermathecal chaetae and thinner cylindrical penial sheaths. A. apeniatus is unique in the genus by having no penis. I. mesoprostatus is distinguishable from congeners by its prostate glands joining middle portion of atria and having concave, cone-shaped cuticular penial sheaths. Twenty-eight species of freshwater oligochaetes have hitherto been recorded from Yunnan Province, including five endemic species from three plateau lakes.

Fridericia dianchiensis sp nov (Enchytraeidae, oligochaeta) from Yunnan Province, China

Fridericia dianchiensis, a new enchytraeid species collected from Yunnan Province, is described here. It is characterized by a combination of the following characters: 1) lateral bundles containing maximum 3 chaetae; 2) esophageal appendages with 3-4 simple, terminal branches; 3) dorsal vessel originating in XX-XXIII; 4) sub-neural glands absent; 5) seminal vesicle large, occupying two segments; 6)clitellum girdle-shaped or gland cells absent between bursal slits and pre-middle ventrally; 7) coelomocytes without refractile vesicles, 8) spermatheca without diverticula and both ampullae broadly united; and 9) long spermathecal ectal duct without gland at the orifice.

Redescription of Nais inflata Liang (Annelida : Oligochaeta) from China

Nais inflata Liang 1963 is redescribed on the basis of fully mature specimens collected from several localities of the Yangtze River and Yellow River. The observations and redescription resulting from the study of mature specimens supports N. inflata as a valid species. This species is characterized by a thickened body, the ventral crotchets all of the same type with distal tooth usually longer than proximal one, a greater number of dorsal chaetae per bundle, stout dorsal needles with equal minute bifid teeth (<1 mu m), presence of penial chaetae, and prostate glands on vasa deferentia instead of atria. The species is most closely related to N. communis Piguet, 1906 and N. variabilis Piguet, 1906. A table comparing allied species is provided.

Potamothrix scleropenis sp nov (Oligochaeta : Tubificidae) from Fuxian Lake, the deepest lake in southwest China

Potamothrix scleropenis sp. nov. (Tubificidae: Tubificinae) is described from the profundal zone (74 m) of Fuxian Lake, the deepest lake (up to 155 m) on the Yunnan-Guizhou Plateau in China. The new species is assigned to Potamothrix because of its short vasa deferentia. and its tubular atria without ejaculatory ducts and prostate glands. It differs from congeners by its cuticularized penis sheaths; bifurcated, strongly curved spermathecal chaetae; bifurcated lower prongs of bifids; and feathered hairs. P scleropenis appears closely related to P cekanovskajae Finogenova, 1972 and P tudoranceai porka, 1994, since all the three species have homogeneous atria without prostate glands.

Bryodrilus fuscistriatus, a new enchytraeid species (Oligochaeta : Enchytraeidae) from northeastern China

Bryodrilus fuscistriatus, a new enchytraeid species from Mt. Changbaishan, Jilin Province, north-eastern China, is described. It is characterized by brown epidermal glands, 7 pairs of preclitellar nephridia, poorly-developed clitellar glands, spermatheca with 2 sessile globular diverticula, and a long sperm funnel with a very broad collar. It is similar to the Alaskan B. tunicatus Dozsa-Farkas & Christensen, 2002 in possessing two diverticula in the spermathecal ampulla and the origin of the dorsal vessel, and the Chinese B. longifistulatus and B. macrotheca Xie et al., 2000c in body size, long sperm funnel and undeveloped clitellar glands, but it differs from B. tunicatus by the presence of brown-striped epidermal gland cells in III-V, a poorly-elevated clitellum, the absence of copulatory glands in XIII-XIV, the regular outline of coelomocytes, and 7 pairs of preclitellar nephridia; from B. longifistulatus and B. macrotheca by the shape of spermatheca, the color of epidermal gland cells, the position of the first pair nephridia, and the origin of dorsal vessel.