The rockefeller university

Recent evidence suggests a critical role for the intracellular signaling protein glycogen synthase kinase-3 (GSK3) in hyperactivity associated with dopaminergic transmission. Here, we investigated whether activation of GSK3 is necessary for the expression of behaviors specifically produced by dopamine D1 receptor activation. To assess the role of GSK3 in dopamine D1 receptor-induced hyperactivity, mice were pretreated with the selective GSK3 inhibitor SB 216763 (0.25-7.5 mg/kg, i.p.) or its vehicle prior to administration of the dopamine D1 receptor full-agonist SKF-82958 (1.0 mg/kg, i.p.) or saline control. Inhibition of GSK3 via SB 216763 dose-dependently reduced ambulatory and stereotypic activity produced by SKF-82958. These data implicate a role for GSK3 in the behavioral manifestations associated with dopamine D1 receptor activation. (C) 2010 Elsevier Inc. All rights reserved.

Terminal differentiation of male germ cells in Drosophila and mammals requires extensive cytoarchitectural remodeling, the elimination of many organelles, and a large reduction in cell volume. The associated process, termed spermatid individualization, is facilitated by the apoptotic machinery, including caspases, but does not result in cell death. From a screen for genes defective in caspase activation in this system, we isolated a novel F-box protein, which we termed Nutcracker, that is strictly required for caspase activation and sperm differentiation. Nutcracker interacts through its F-box domain with members of a Cullin-1-based ubiquitin ligase complex (SCF): Cullin-1 and SkpA. This ubiquitin ligase does not regulate the stability of the caspase inhibitors DIAP1 and DIAP2, but physically binds Bruce, a BIR-containing giant protein involved in apoptosis regulation. Furthermore, nutcracker mutants disrupt proteasome activity without affecting their distribution. These findings define a new SCF complex required for caspase activation during sperm differentiation and highlight the role of regulated proteolysis during this process.

Women and people of color continue to be underrepresented among biomedical researchers to an alarming degree. Research interest and subsequent productivity have been shown to be affected by the research training environment through the mediating effects of research self-efficacy. This article presents the findings of a study to determine whether a short-term research training program coupled with an efficacy enhancing intervention for novice female biomedical scientists of diverse racial backgrounds would increase their research self-efficacy beliefs. Forty-three female biomedical scientists were randomized into a control or intervention group and 15 men participated as a control group. Research self-efficacy significantly increased for women who participated in the self-efficacy intervention workshop. Research self-efficacy within each group also significantly increased following the short-term research training program, but cross-group comparisons were not significant. These findings suggest that educational interventions that target sources of self-efficacy and provide domain-specific learning experiences are effective at increasing research self-efficacy for women and men. Further studies are needed to determine the longitudinal outcomes of this effort.

Vaccinations in medicine are commonly administered through the skin. Therefore, the vaccine is immunologically processed by antigen-presenting cells of the skin. There is recent evidence that the clinically less often used intradermal route is effective; in cases even superior to the conventional subcutaneous or intramuscular route. Professional antigen-presenting cells of the skin comprise epidermal Langerhans cells (CD207/langerin(+)), dermal langerin(-) and dermal langerin(+) dendritic cells (DCs). In human skin, langerin(-) dermal DCs can be further subdivided on the basis of their reciprocal CD1a and CD14 expression. The relative contributions of these subsets to the generation of immunity or tolerance are still unclear. Langerhans cells in human skin seem to be specialized for induction of cytotoxic T lymphocytes. Likewise, mouse Langerhans cells are capable of cross-presentation and of protecting against experimental tumours. It is desirable to harness these properties for immunotherapy. A promising strategy to dramatically improve the outcome of vaccinations is 'antigen targeting'. Thereby, the vaccine is delivered directly and selectively to defined types of skin DCs. Targeting is achieved by means of coupling antigen to antibodies that recognize cell surface receptors on DCs. This approach is being widely explored. Little is known, however, about the events that take place in the skin and the DCs subsets involved therein. This topic will be discussed in this article. Immunology and Cell Biology (2010) 88, 424-430; doi: 10.1038/icb.2010.39; published online 6 April 2010

We report the application of a far-from-equilibrium statistical-mechanical theory to a nontrivial system with Newtonian interactions in continuous boundary-driven flow. By numerically time stepping the force-balance equations of a one-dimensional model fluid we measure occupancies and transition rates in simulation. The high-shear-rate simulation data reproduce the predicted invariant quantities, thus supporting the theory that a class of nonequilibrium steady states of matter, namely, sheared complex fluids, is amenable to statistical treatment from first principles.

Elucidating virus-host interactions responsible for HIV-1 transmission is important for advancing HIV-1 prevention strategies. To this end, single genome amplification (SGA) and sequencing of HIV-1 within the context of a model of random virus evolution has made possible for the first time an unambiguous identification of transmitted/founder viruses and a precise estimation of their numbers. Here, we applied this approach to HIV-1 env analyses in a cohort of acutely infected men who have sex with men (MSM) and found that a high proportion (10 of 28; 36%) had been productively infected by more than one virus. In subjects with multivariant transmission, the minimum number of transmitted viruses ranged from 2 to 10 with viral recombination leading to rapid and extensive genetic shuffling among virus lineages. A combined analysis of these results, together with recently published findings based on identical SGA methods in largely heterosexual (HSX) cohorts, revealed a significantly higher frequency of multivariant transmission in MSM than in HSX [19 of 50 subjects (38%) versus 34 of 175 subjects (19%); Fisher's exact p = 0.008]. To further evaluate the SGA strategy for identifying transmitted/founder viruses, we analyzed 239 overlapping 5' and 3' half genome or env-only sequences from plasma viral RNA (vRNA) and blood mononuclear cell DNA in an MSM subject who had a particularly well-documented virus exposure history 3 -6 days before symptom onset and 14-17 days before peak plasma viremia (47,600,000 vRNA molecules/ml). All 239 sequences coalesced to a single transmitted/ founder virus genome in a time frame consistent with the clinical history, and a molecular clone of this genome encoded replication competent virus in accord with model predictions. Higher multiplicity of HIV-1 infection in MSM compared with HSX is consistent with the demonstrably higher epidemiological risk of virus acquisition in MSM and could indicate a greater challenge for HIV-1 vaccines than previously recognized.

Most HIV-infected individuals develop antibodies to the gp120 and gp41 components of the viral spike; however, only a fraction of these individuals mount a broadly neutralizing serum response against HIV. We have cloned anti-HIV antibodies from the memory B-cell compartment of six individuals with variable viral loads and high titers of broadly neutralizing antibodies. Here, we report on the features of the anti-gp41 response in these patients. Competition experiments with previously characterized antibodies targeting defined epitopes on the gp41 ectodomain showed antibodies directed against the "immunodominant region" (cluster I), the carboxy-terminal heptad repeat (cluster II), and the membrane-proximal external region (cluster IV). On the other hand, antibodies directed against the amino-terminal part of the molecule, including the fusion peptide, polar region, and the N-terminal heptad repeat, were not detected. When all patients' data were combined, unique B-cell clones targeting cluster I, II, and IV accounted for 32%, 49%, and 53% of all anti-gp41-reactive B cells, respectively; therefore, no single region was truly immunodominant. Finally, although we found no new neutralizing epitopes or HIV-1-neutralizing activity by any of the gp41 antibodies at concentrations of up to 50 mu g/ml, high concentrations of 7 out of 15 anti-cluster I antibodies neutralized tier 2 viruses.

This article starts in Part I with a simple example of two biochemical reaction networks that are indistinguishable at the macroscopic level but are different at the molecular level and are shown to have significantly different kinetic properties. So, if one completely ignores the fact that reactions advance in discrete steps at the molecular level, then one can fail to distinguish between networks with widely different kinetics. In part II biochemical reaction networks are treated in a general way to discover what property of a network, only seen at the molecular level, affects its kinetics. It is shown that every such network has a unique torsion group which can be described numerically and readily determined by a programmable computation. If the group is found to be the singleton {0} (as is most often the case in practice), then the network is said to be torsion-free and its kinetic properties unaffected by ignoring its discrete character. A chemical reaction network has to be represented algebraically to calculate its torsion group. If the network is to be understood only at the macroscopic level, it can be placed in the context of real vector spaces, but to recognize its discrete character and its torsion group, each vector space is replaced by a discrete subset of that space, where each molecule can be recognized as a distinct and indivisible entity. Next, the process of calculating a torsion group is shown in several cases, including the example in part I. In this particular case it is shown to have the torsion group with 2 elements, reflecting the fact that the substrate molecules become product molecules 2 at a time, with the result that the overall macroscopic reaction is R a double dagger" T, whereas at the molecular level it is 2R a double dagger" 2T. In general, however, the torsion group of a biochemical reaction network can be any finite additive group, which is a property of the network that can only be seen at the molecular level. Finally, this fact is demonstrated by showing how to construct a hypothetical, but plausible, biochemical reaction network that has any given finite additive group as its torsion group.

Rhodopsin is a prototypical heptahelical family A G-protein-coupled receptor (GPCR) responsible for dim-light vision(1,2). Light isomerizes rhodopsin's retinal chromophore and triggers concerted movements of transmembrane helices, including an outward tilting of helix 6 (H6) and a smaller movement of H5, to create a site for G-protein binding and activation(3,4). However, the precise temporal sequence and mechanism underlying these helix rearrangements is unclear. We used site-directed non-natural amino acid mutagenesis to engineer rhodopsin with p-azido-L-phenylalanine residues incorporated at selected sites(5), and monitored the azido vibrational signatures using infrared spectroscopy as rhodopsin proceeded along its activation pathway. Here we report significant changes in electrostatic environments of the azido probes even in the inactive photoproduct Meta I, well before the active receptor state was formed. These early changes suggest a significant rotation of H6 and movement of the cytoplasmic part of H5 away from H3. Subsequently, a large outward tilt of H6 leads to opening of the cytoplasmic surface to form the active receptor photoproduct Meta II(3). Thus, our results reveal early conformational changes that precede larger rigid-body helix movements, and provide a basis to interpret recent GPCR crystal structures(6,7) and to understand conformational sub-states observed during the activation of other GPCRs(8).