Inhibition of Simian Virus 40 replication by targeting the molecular chaperone function and ATPase activity of T antigen

Polyomaviruses such as BK virus and JC virus have been linked to several diseases, but treatments that thwart their propagation are limited in part because of slow growth and cumbersome culturing conditions. In contrast, the replication of one member of this family, Simian Virus 40 (SV40), is robust and has been well-characterized. SV40 replication requires two domains within the viral-encoded large tumor antigen (TAg): The ATPase domain and the N-terminal J domain, which stimulates the ATPase activity of the Hsp70 chaperone. To assess whether inhibitors of polyomavirus replication could be identified, we examined a recently described library of small molecules, some of which inhibit chaperone function. One compound, MAL2-11B, inhibited both TAg's endogenous ATPase activity and the TAg-mediated activation of Hsp70. MAL2-11B also reduced SV40 propagation in plaque assays and compromised DNA replication in cell culture and in vitro. Furthermore, the compound significantly reduced the growth of BK virus in a human kidney cell line. These data indicate that pharmacological inhibition of TAg's chaperone and ATPase activities may provide a route to combat polyomavirus-mediated disease. © 2009 Elsevier B.V. All rights reserved

A Millimeter-Wave Klystron Upconverter With a Higher Order Mode Output Cavity

Manufacturing of klystrons in the millimeter-wave frequency range is challenging due to the small size of the cavities and the ratio of the maximum gap voltage to the beam energy. The small dimensions also make difficult to produce devices with the output power required by a number of applications at millimeter wave, such as communications and spectroscopy. Operating with a higher order mode can be a potential solution, as a larger transverse size structure can be used. Unfortunately, high-order mode cavities have a lower impedance than in fundamental mode. In this paper is proposed a novel solution to overcome the reduced impedance by utilizing an upconverter, where all cavities except the output cavity are designed to work in high-order mode. To demonstrate the effectiveness of the approach, two klystron upconverters were designed. One has six cavities aiming to achieve a maximum output power of $~$90 W at 105 GHz. The second klystron upconverter was a simpler three-cavity structure designed for quick prototype. Millimeter-wave measurements of the three-cavity klystron upconverter are presented

Normalizing glycosphingolipids restores function in CD4+ T cells from lupus patients

Patients with the autoimmune rheumatic disease systemic lupus erythematosus (SLE) have multiple defects in lymphocyte signaling and function that contribute to disease pathogenesis. Such defects could be attributed to alterations in metabolic processes, including abnormal control of lipid biosynthesis pathways. Here, we reveal that CD4+ T cells from SLE patients displayed an altered profile of lipid raft–associated glycosphingolipids (GSLs) compared with that of healthy controls. In particular, lactosylceramide, globotriaosylceramide (Gb3), and monosialotetrahexosylganglioside (GM1) levels were markedly increased. Elevated GSLs in SLE patients were associated with increased expression of liver X receptor β (LXRβ), a nuclear receptor that controls cellular lipid metabolism and trafficking and influences acquired immune responses. Stimulation of CD4+ T cells isolated from healthy donors with synthetic and endogenous LXR agonists promoted GSL expression, which was blocked by an LXR antagonist. Increased GSL expression in CD4+ T cells was associated with intracellular accumulation and accelerated trafficking of GSL, reminiscent of cells from patients with glycolipid storage diseases. Inhibition of GSL biosynthesis in vitro with a clinically approved inhibitor (N-butyldeoxynojirimycin) normalized GSL metabolism, corrected CD4+ T cell signaling and functional defects, and decreased anti-dsDNA antibody production by autologous B cells in SLE patients. Our data demonstrate that lipid metabolism defects contribute to SLE pathogenesis and suggest that targeting GSL biosynthesis restores T cell function in SLE

Reconstituted B cell receptor signaling reveals carbohydrate-dependent mode of activation

Activation of immune cells (but not B cells) with lectins is widely known. We used the structurally defined interaction between influenza hemagglutinin (HA) and its cell surface receptor sialic acid (SA) to identify a B cell receptor (BCR) activation modality that proceeded through non-cognate interactions with antigen. Using a new approach to reconstitute antigen-receptor interactions in a human reporter B cell line, we found that sequence-defined BCRs from the human germline repertoire could be triggered by both complementarity to influenza HA and a separate mode of signaling that relied on multivalent ligation of BCR sialyl-oligosaccharide. The latter suggested a new mechanism for priming naïve B cell responses and manifested as the induction of SA-dependent pan-activation by peripheral blood B cells. BCR crosslinking in the absence of complementarity is a superantigen effect induced by some microbial products to subvert production of antigen-specific immune responses. B cell superantigen activity through affinity for BCR carbohydrate is discussed

Real-time intermembrane force measurements and imaging of lipid domain morphology during hemifusion

Membrane fusion is the core process in membrane trafficking and is essential for cellular transport of proteins and other biomacromolecules. During protein-mediated membrane fusion, membrane proteins are often excluded from the membrane-membrane contact, indicating that local structural transformations in lipid domains play a major role. However, the rearrangements of lipid domains during fusion have not been thoroughly examined. Here using a newly developed Fluorescence Surface Forces Apparatus (FL-SFA), migration of liquid-disordered clusters and depletion of liquid-ordered domains at the membrane-membrane contact are imaged in real time during hemifusion of model lipid membranes, together with simultaneous force-distance and lipid membrane thickness measurements. The load and contact time-dependent hemifusion results show that the domain rearrangements decrease the energy barrier to fusion, illustrating the significance of dynamic domain transformations in membrane fusion processes. Importantly, the FL-SFA can unambiguously correlate interaction forces and in situ imaging in many dynamic interfacial systems.open0