Molecular cloning of the murine IL-1 beta converting enzyme cDNA

IL-1 beta is a potent modulator of immune and inflammatory responses. Murine IL-1 beta is initially synthesized as an inactive 33-kDa pro-molecule that is activated by proteolytic cleavage between Asp-117 and Val-118 to generate the 17-kDa mature IL-1 beta protein. This cleavage is catalyzed by a specific protease that has been designated the IL-1 beta converting enzyme (or IL-1 beta convertase). We have used a human IL-1 beta convertase cDNA to isolate murine convertase cDNA from a WEHI-3 library. These cDNA predicted that the murine convertase is a 402-residue protein. Overall, the murine convertase showed 71% nucleotide and 62% predicted amino acid sequence identity with the human convertase. Southern blot analysis of interspecific backcross mice indicated that the murine IL-1 beta convertase is encoded by a single copy gene located on murine chromosome 9. The murine convertase showed broad constitutive expression, being detected in mononuclear phagocyte and T lymphocyte cell lines as well as in spleen, heart, brain, and adrenal glands. The expression of the murine convertase in mononuclear phagocytes was up-regulated by treatment with LPS or rIFN-gamma. These studies establish that the IL-1 beta convertase is an evolutionarily conserved, widely expressed enzyme that can be regulated at a pretranslational level.     

Discovery of selective, small-molecule inhibitors of RNA complexes--I. The Tat protein/TAR RNA complexes required for HIV-1 transcription

We have developed a therapeutic program focusing on the inhibition of a human immunodeficiency virus-1 specific protein-RNA interaction. This program begins with a search for small organic molecules that would interfere with the binding of Tat protein to TAR RNA. The methodologies chosen to study the HIV-1 Tat-TAR interaction and inhibition include gel mobility shift assays, scintillation proximity assays, filtration assays, and mass spectrometry. These methods helped establish in vitro high-throughput screening assays which rapidly identified Tat-TAR inhibitors from our corporate compound library. Tat-activated reporter gene assays were then used to investigate the cellular activities of the Tat-TAR inhibitors. The cellular activity, selectivity, and toxicity data for select Tat-TAR inhibitors were determined. Evaluation of both the cellular data and the Tat-TAR inhibition results led to further testing in anti-HIV-1 infection assays.     

Ag nanotubes and Ag/AgCl electrodes in nanoporous membranes

Miniaturization of the entire experimental setup is a key requirement for widespread application of nanodevices. For nanopore biosensing, integrating electrodes onto the nanopore membrane and controlling the pore length is important for reducing the complexity and improving the sensitivity of the system. Here we present a method to achieve these goals, which relies on electroless plating to produce Ag nanotubes in track-etched polymer nanopore templates. By plating from one side only, we create a conductive nanotube that does not span the full length of the pore, and thus can act as a nanoelectrode located inside the nanopore. To give optimal electrochemical behavior for sensing, we coat the Ag nanotube with a layer of AgCl. We characterize the behavior of this nanoelectrode by measuring its current-voltage response and find that, in most cases, the response is asymmetric. The plated nanopores have initial diameters between 100 and 300?nm, thus a range suitable for detection of viruses.     

Standards for diagnosis and management of invasive breast carcinoma. American College of Radiology. American College of Surgeons. College of American Pathologists. Society of Surgical Oncology

Because knowledge has advanced in several fields related to the treatment of early breast cancer, revising the landmark 1992 standards for breast-conservation treatment by these four organizations is appropriate. The current report reviews and summarizes the literature and describes the selection and evaluation of patients, the technical aspects of surgical treatment and irradiation, follow-up care, and areas for further research.     

The nematode degenerin UNC-105 forms ion channels that are activated by degeneration- or hypercontraction-causing mutations

Nematode degenerins have been implicated in touch sensitivity and other forms of mechanosensation. Certain mutations in several degenerin genes cause the swelling, vacuolation, and death of neurons, and other mutations in the muscle degenerin gene unc-105 cause hypercontraction. Here, we confirm that unc-105 encodes an ion channel and show that it is constitutively active when mutated. These mutations disrupt different regions of the channel and have different effects on its gating. The UNC-105 channels are permeable to small monovalent cations but show voltage-dependent block by Ca2+ and Mg2+. Amiloride also produces voltage-dependent block, consistent with a single binding site 65% into the electric field. Mammalian cells expressing the mutant channels accumulate membranous whorls and multicompartment vacuoles, hallmarks of degenerin-induced cell death across species.