Evidence of both extra- and intracellular cysteine targets of protein modification for activation of RET kinase

By use of a specifically sulfhydryl group-reactive chemical, 1,4-butanediyl-bismethanethiosulfonate (BMTS), we studied the localization of oxidative stress-responsive target cysteines for activation of a receptor-type protein tyrosine kinase, c-RET. The chemical, which reacted with RET proteins on the cell surface for sulfhydryl-linked aggregation, induced autophosphorylation and activation of RET kinase. When extracellular domain-deleted RET mutant (RET-PTC-1) cells were exposed to BMTS, neither the molecular status nor the activity of the enzyme was affected, suggesting that the target cysteines of BMTS to which cells were exposed for reaction are located in the cysteine-rich region of the extracellular domain of RET kinase. Despite this result, the exposure of a subcellular form of c-RET or RET-PTC-1 kinase isolated by immunoprecipitation to BMTS did induce activation of the enzyme. These results suggest that cysteines in both the extracellular and the intracellular domains of RET can work as target sites of accessible BMTS and possibly other oxidative elements for structural modification and activation of RET kinase.     

Reproductive toxicity and infertility effect of Ferula hornonis extracts in mice

The anti-fertility, anti-implantation, and ovarian histological alterations of the ethanolic extract of Ferula hormonis have been investigated in female mice. The intragastric application of 3 mg/kg per day of such extract for 6 weeks resulted in a significant reduction in female mice fertility. Furthermore, it caused a decrease in the number of mated females, the total number of implantations, and the number of viable fetuses. These changes were also associated with ovarian atrophy and a concomitant increase in the connective tissue. The ova showed degeneration while most of the ovarian follicles suffered follicular atresia.     

Lymphocide: cytokines and the control of lymphoid homeostasis

In a human, about 10(11) excess peripheral lymphocytes die every day. This death process maintains a constant lymphocyte population size in the face of a continuous influx of new lymphocytes and the homeostatic proliferation of old ones. Death is triggered when a lymphocyte fails to acquire signals from survival factors, the availability of which, therefore, determines the size of the pool of lymphocytes. A lymphocyte acquires survival signals through receptors for cytokines, antigens, hormones and probably other extracellular factors. Here, we discuss current concepts of the intracellular signalling pathways for survival versus death that establish cytokine-regulated lymphocyte homeostasis.     

The discovery net system for high throughput bioinformatics

MOTIVATION: Bioinformatics requires Grid technologies and protocols to build high performance applications without focusing on the low level detail of how the individual Grid components operate. RESULTS: The Discovery Net system is a middleware that allows service developers to integrate tools based on existing and emerging Grid standards such as web services. Once integrated, these tools can be used to compose reusable workflows using these services that can later be deployed as new services for others to use. Using the Discovery Net system and a range of different bioinformatics tools, we built a Grid based application for Genome Annotation. This includes workflows for automatic nucleotide annotation, annotation of predicted proteins and text analysis based on metabolic profiles and text analysis.     

Dexiotrichides pangi n. sp. (protozoa,ciliophora, scuticociliatia), a new marine ciliate from the north China sea

The morphology, infraciliature, and silverline system of a new marine scuticociliate, Dexiotrichides pangi n. sp. were investigated. The new species is characterized by: size about 45-65 x 20-25 microm in vivo with kidney-like body shape and obliquely truncated semicircle-shaped apical plate; cytostome at bottom of conspicuously depressed oral cavity, which is located at the cell equatorial level; paroral membrane extending anteriorly to membranelle 3; scutica multi-rowed; 33-38 somatic kineties; contractile vacuole near ventral side and subcaudally positioned, opening at posterior end of somatic kinety 3; one oval macronucleus and one small micronucleus; caudal cilium positioned in a small pouch; marine habitat. Based on the data obtained, an improved diagnosis for the genus Dexiotrichides is suggested: body with circular cross-section and conspicuous cilia-free apical plate; buccal cavity conspicuously depressed with cytostome located near or at equatorial level; three membranelles transversely orientated each with 2-3 rows; paroral membrane zigzaging structure, extending to about half of the length of buccal field; multi-rowed scutica; somatic kinety one strongly shortened and terminating anteriorly at posterior end of buccal field; basal bodies in equatorial region arranged usually in circular pattern, while in the anterior portion of somatic kinety 2, basal bodies characteristically in pairs and separated from the posterior part of kinety 2; one caudal cilium.     

Ca2+-calmodulin-dependent protein kinase II potentiates store-operated Ca2+ current

A rise in intracellular Ca2+ (Ca2+i) mediates various cellular functions ranging from fertilization to gene expression. A ubiquitous Ca2+ influx pathway that contributes significantly to the generation of Ca2+i signals, especially in non-excitable cells, is store-operated Ca2+ entry (SOCE). Consequently, the modulation of SOCE current affects Ca2+i dynamics and thus the ensuing cellular response. Therefore, it is important to define the mechanisms that regulate SOCE. Here we show that a rise in Ca2+i potentiates SOCE. This potentiation is mediated by Ca2+-calmodulin-dependent protein kinase II (CaMKII), because inhibition of endogenous CaMKII activity abrogates Ca2+i-mediated SOCE potentiation and expression of constitutively active CaMKII potentiates SOCE current independently of Ca2+i. Moreover, we present evidence that CaMKII potentiates SOCE by altering SOCE channel gating. The regulation of SOCE by CaMKII defines a novel modulatory mechanism of SOCE with important physiological consequences.     

Tissue- and stressor-specific differential expression of two hsc70 genes in carp

Two genes expressing 70 kDa heat shock proteins were identified in Cyprinus carpio. The sequence similarities and the intron-interrupted structure of the coding regions indicate that carp Hsc70-1 and Hsc70-2 belong to the Hsp70 cognate subfamily. The expressions of the two hsc70 genes were followed by semi-quantitative RT-PCR. Both genes are expressed under unstressed conditions in a characteristic tissue-specific manner. Inducibility of the response to elevated temperature, cold shock, and Cd treatment was investigated in the liver and muscle, in whole-animal experiments. Both genes were insensitive to or only weakly induced by the stressors, with two exceptions: Cd treatment resulted in an 11-13-fold enhanced induction of hsc70-1 in the liver and cold shock enhanced induction of hsc70-2 in the muscle by 7.5-10-fold.     

Caspase-3 can be pseudo-activated by a Ca2+-dependent proteolysis at a non-canonical site

We have shown previously that calcium could trigger nuclear fragmentation, which was associated with a caspase 3 (C3)-like activity [Juin, P., Pelletier, M., Oliver, L., Tremblais, K., Gregoire, M., Meflah, K. and Vallette, F.M. (1998) Induction of a caspase-3-like activity by calcium in normal cytosolic extracts triggers nuclear apoptosis in a cell-free system. J. Biol. Chem. 273, 17559]. Here, we report that this activation is associated with a non-canonical truncation of C3, which induces a weak DEVDase activity. The cleavage of C3 via calcium-dependent proteolysis is independent of caspase 9; lysate exposure to calcium prevents further cleavage and activation by the cytochrome c and dATP pathway. Altogether, our data suggest that calcium could favour a necrotic mechanism by inducing the generation of a form of C3 insensitive to mitochondrial activation.     

Distinct domains control the addressing and the insertion of Bax into mitochondria

The translocation of Bax from the cytosol into the mitochondrial outer membrane is a central event during apoptosis. We report that beyond the addressing step, which involves its first alpha-helix (halpha1), the helices alpha5 and alpha6 (halpha5alpha6) are responsible for the insertion of Bax into mitochondrial outer membrane bilayer. The translocation of Bax to mitochondria is associated with specific changes in the conformation of the protein that are under the control of two prolines: Pro-13, which controls the unfolding of halpha1, and Pro-168, a proline located immediately before the hydrophobic carboxyl-terminal end (i.e. helix alpha9, halpha9), which controls the disclosure of halpha5alpha6. An additional step, the disruption of an electrostatic bond formed between Asp-33 (halpha1) and Lys-64 (BH3), allows the mitochondria addressing of Bax. We conclude that, although the intramolecular interactions of halpha1 with the BH3 region control the addressing of Bax to mitochondria, the Pro-168 is involved in the control of its membrane insertion through halpha5alpha6.