Structure of astacin and implications for activation of astacins and zinc-ligation of collagenases.

Astacin, a digestive zinc-endopeptidase from the crayfish Astacus astacus L., is the prototype for the 'astacin family', which includes mammalian metallo-endopeptidases and developmentally regulated proteins of man, fruitfly, frog and sea urchin. Here we report the X-ray crystal structure of astacin, which reveals a deep active-site cleft, with the zinc at its bottom ligated by three histidines, a water molecule and a more remote tyrosine. The third histidine (His 102) forms part of a consensus sequence, shared not only by the members of the astacin family, but also by otherwise sequentially unrelated proteinases, such as vertebrate collagenases. It may therefore represent the elusive 'third' zinc ligand in these enzymes. The amino terminus of astacin is buried forming an internal salt-bridge with Glu 103, adjacent to His 102. Astacin pro-forms extended at the N terminus, as observed for some 'latent' mammalian astacin homologues, did not exhibit this 'active' conformation, indicating an activation mechanism reminiscent of trypsin-like serine proteinases.     

Choroidal and iris angioarchitecture of the newt: A scanning electron-microscopic study of vascular corrosion casts

The corrosion cast technique provided for the first time an excellent three-dimensional visualization of the vascular pattern of the choroid and iris in the newt eye. The results show the presence of a single arterial afference to the choroidal and iris capillaries: the ophthalmic artery is the origin fo both ciliary arteries and the long posterior ciliary artery. Slightly behind the equatorial circumference of the eyeball the venous drainage consists of a single vessel on the dorsal side and two distinct vessels on the ventral one. It receives blood from both iris and choroid. The surface of the plastic endocasts shows some details of fine luminal structures of the endothelial cells. Shallow depressions may be regarded as imprints of endothelial cell nuclei, and they are distinctly different for arteries and capillaries. The angioarchitecture of the newt eye differs from that of brain in that hairpin-shaped capillary loops are not observed at all.     

Exocytotic fusion is activated by Rab3a peptides.

Studies of intracellular traffic in yeast and mammalian systems have implicated members of the Rab family of small GTP-binding proteins as regulators of membrane fusion. We have used the patch clamp technique to measure exocytotic fusion events directly and investigate the role of GTP-binding proteins in regulating exocytosis in mast cells. Intracellular perfusion of mast cells with GTP-gamma S is sufficient to trigger complete exocytotic degranulation in the absence of other intracellular messengers. Here we show that GTP is a potent inhibitor of GTP-gamma S-induced degranulation, indicating that sustained activation of a GTP-binding protein is sufficient for membrane fusion. We have found that synthetic oligopeptides, corresponding to part of the effector domain of Rab3a, stimulate complete exocytotic degranulation, similar to that induced by GTP-gamma S. The response is selective for Rab3a sequence and is strictly dependent on Mg2+ and ATP. This suggests that sustained activation of a Rab3 protein causes exocytotic fusion. The peptide response can be accelerated by GDP-beta S, suggesting that Rab3a peptides compete with endogenous Rab3 proteins for a binding site on a target effector protein, which causes fusion on activation.     

How do Shp2 mutations that oppositely influence its biochemical activity result in syndromes with overlapping symptoms?

Activating and inactivating mutations of SHP-2 are responsible, respectively, for the Noonan (NS) and the LEOPARD (LS) syndromes. Clinically, these developmental disorders overlap greatly, resulting in the apparent paradox of similar diseases caused by mutations that oppositely influence SHP-2 phosphatase activity. While the mechanisms remain unclear, recent functional analysis of SHP-2, along with the identification of other genes involved in NS and in other related syndromes (neurofibromatosis-1, Costello and cardio-facio-cutaneous syndromes), strongly suggest that Ras/MAPK represents the major signaling pathway deregulated by SHP-2 mutants. We discuss the idea that, with the exception of LS mutations that have been shown to exert a dominant negative effect, all disease-causing mutations involved in Ras/MAPK-mediated signaling, including SHP-2, might lead to enhanced MAPK activation. This suggests that a narrow range of MAPK signaling is required for appropriate development. We also discuss the possibility that LS mutations may not simply exhibit dominant negative activity. Received 30 November 2006; received after revision 8 February 2007; accepted 13 March 2007     

Targeted inhibition of Livin resensitizes renal cancer cells towards apoptosis

Cancer cells are typically characterized by apoptosis deficiency. In order to investigate a possible role for the anti-apoptotic livin gene in renal cell cancer (RCC), we analyzed its expression in tumor tissue samples and in RCC-derived cell lines. In addition, we studied the contribution of livin to the apoptotic resistance of RCC cells by RNA interference (RNAi). Livin gene expression was detected in a significant portion of RCC tumor tissue specimens (13/14, 92.9%) and tumor-derived cell lines (12/15, 80.0%). Moreover, targeted inhibition of livin by RNAi markedly sensitized RCC cells towards proapoptotic stimuli, such as UV irradiation or the chemotherapeutic drugs etoposide, 5-fluorouracil, and vinblastine. These effects were specific for livin expressing tumor cells. We conclude that livin can contribute significantly to the apoptosis resistance of RCC cells. Targeted inhibition of livin could represent a novel therapeutic strategy to increase the sensitivity of renal cancers towards pro-apoptotic agents. Received 30 November 2006; received after revision 22 February 2007; accepted 20 March 2007     

Common Molecular Mechanisms of Mammary Gland Development and Breast Cancer

During its lifetime, the mammary gland undergoes many phases of development and differentiation. Much of this occurs during puberty, when the ductal epithelium expands by branching morphogenesis, invading the surrounding fat pad to form an organised mammary tree. Throughout its existence, the epithelium will go through several cycles of proliferation and cell death during pregnancy, lactation and involution. Many of the signalling mechanisms which control the initial invasion of the fat pad by the epithelium, and regulate its continuing plasticity, can be harnessed or corrupted by tumour cells in order to support their aberrant growth and progression towards invasion. This is true not just for the epithelial cells themselves but also for cells in the surrounding microenvironment, including fibroblasts, macrophages and adipocytes. This review examines the complex web of signalling and adhesion interactions controlling branching morphogenesis, and how their alteration can promote malignancy. Current in vivo and in vitro mammary gland models are also discussed. (Part of a Multi-author Review)     

Nanostructured Functional Thermoplastic Polymeric Materials Based on the Molecular Control of the Blending

1 Results The development of the concepts of nanotechnology has given an important impact on the design of new polymer based materials which are in most cases characterized by a multiphase morphology. When at least one phase has nanometric dimension(s) the system can be considered as a nanocomposite where the interface is not only determining for the adhesion but also may play a role in some bulk properties. Indeed in nanostructured multiphase solids the interface is significant as a bulk component. The...     

The Three-component One-pot Synthesis of 4,6-Diarylpyrimidin-2(1h)-ones and 9-Phenyl-8-oxa-10,12-diaza-tricyclo[7.3.1.0~(2,7)]trideca-2(7),3,5-trien-11-one

1 Results Pyrimidinones (PMs) are a class of important heterocycles which have been well documented throughout the literature due to their biological importance. They exhibit a wide range of pharmaceutical and therapeutic properties[1].A rapid and efficient one-pot method for the synthesis of 4,6-diarylpyrimidin-2(1H)- ones and related heterocycles is described.The condensation of acetophenone derivatives,aldehydes and urea in the presence of sulfamic acid was employed to synthesize a variety of pyrimid...     

Notizie preliminari sull'azione fitodinamica dell'acido difenililacetico sulla pianta del tabacco

Summary The application of diphenylilacetic acid has shown favorable effects on the growth of tobacco plants. Solutions containing 10 p.p.m. and 20 p.p.m. produce, in weight, an almost double growth in treated plants in comparison with the controls. Diphenylilacetic acid can be considered as a growth-promoting substance. The data are not definite.