Glutamatergic plasticity by synaptic delivery of GluR-B(long)-containing AMPA receptors

Activity-driven delivery of AMPA receptors is proposed to mediate glutamatergic synaptic plasticity, both during development and learning. In hippocampal CA1 principal neurons, such trafficking is primarily mediated by the abundant GluR-A subunit. We now report a study of GluR-B(long), a C-terminal splice variant of the GluR-B subunit. GluR-B(long) synaptic delivery is regulated by two forms of activity. Spontaneous synaptic activity-driven GluR-B(long) transport maintains one-third of the steady-state AMPA receptor-mediated responses, while GluR-B(long) delivery following the induction of LTP is responsible for approximately 50% of the resulting potentiation at the hippocampal CA3 to CA1 synapses at the time of GluR-B(long) peak expression-the second postnatal week. Trafficking of GluR-B(long)-containing receptors thus mediates a GluR-A-independent form of glutamatergic synaptic plasticity in the juvenile hippocampus.     

The function of GPI-anchored proteins in T cell development, activation and regulation of homeostasis

Many glycosylphosphatidyl-inositol-anchored proteins (GPI-AP) are expressed on T lymphocytes. Ligand or mAb-mediated aggregation of all GPI-AP tested to date results in the initiation of signal transduction pathways via the activation of src family protein tyrosine kinases. Src family kinases co-localise with GPI-AP in specialised sub-domains of the plasma membrane, referred to as detergent insoluble membrane microdomains (DIGS), which are thought to function as signalling platforms. GPI-AP may play a role in the regulation of T cell clonal expansion and effector functions at multiple levels, including the initiation of T cell activation through the antigen receptor complex, the regulation of ongoing responses supported by persisting antigen, as well as proliferative responses to the major T cell growth factor, IL-2. Evidence supporting the role of GPI-AP in the regulation of T cell development, activation and homeostasis is discussed, as well as insights provided by studies in humans and mice lacking GPI-AP.     

Q vectors, bicistronic retroviral vectors for gene transfer

We have developed a retroviral vector that incorporates unique features of some previously described vectors. This vector includes: 3' long terminal repeats (LTRs) of the self-inactivating class; a 5' LTR that is a hybrid of the cytomegalovirus (CMV) enhancer and the mouse sarcoma virus promoter; an internal CMV immediate early region promoter to drive expression of the transduced gene and the neomycin phosphotransferase selectable marker; an expanded multiple cloning site and an internal ribosome entry site. An SV40 ori was introduced into the vector backbone to promote high copy number replication in packaging cell lines that express the SV40 large T antigen. We demonstrate that these retroviral constructs, designated Q vectors, can be used in applications where high viral titers and high level stable or transient gene expression are desirable.     

The 2'-O-ribose methyltransferase for cap 1 of spliced leader RNA and U1 small nuclear RNA in Trypanosoma brucei

mRNA cap 1 2'-O-ribose methylation is a widespread modification that is implicated in processing, trafficking, and translational control in eukaryotic systems. The eukaryotic enzyme has yet to be identified. In kinetoplastid flagellates trans-splicing of spliced leader (SL) to polycistronic precursors conveys a hypermethylated cap 4, including a cap 0 m7G and seven additional methylations on the first 4 nucleotides, to all nuclear mRNAs. We report the first eukaryotic cap 1 2'-O-ribose methyltransferase, TbMTr1, a member of a conserved family of viral and eukaryotic enzymes. Recombinant TbMTr1 methylates the ribose of the first nucleotide of an m7G-capped substrate. Knockdowns and null mutants of TbMTr1 in Trypanosoma brucei grow normally, with loss of 2'-O-ribose methylation at cap 1 on substrate SL RNA and U1 small nuclear RNA. TbMTr1-null cells have an accumulation of cap 0 substrate without further methylation, while spliced mRNA is modified efficiently at position 4 in the absence of 2'-O-ribose methylation at position 1; downstream cap 4 methylations are independent of cap 1. Based on TbMTr1-green fluorescent protein localization, 2'-O-ribose methylation at position 1 occurs in the nucleus. Accumulation of 3'-extended SL RNA substrate indicates a delay in processing and suggests a synergistic role for cap 1 in maturation.     

Comparative Metabolism of Free‐living Bodo saltans and Parasitic Trypanosomatids

Comparison of the genomes of free‐living Bodo saltans and those of parasitic trypanosomatids reveals that the transition from a free‐living to a parasitic life style has resulted in the loss of approximately 50% of protein‐coding genes. Despite this dramatic reduction in genome size, B. saltans and trypanosomatids still share a significant number of common metabolic traits: glycosomes; a unique set of the pyrimidine biosynthetic pathway genes; an ATP‐PFK which is homologous to the bacterial PP_i‐PFKs rather than to the canonical eukaryotic ATP‐PFKs; an alternative oxidase; three phosphoglycerate kinases and two GAPDH isoenzymes; a pyruvate kinase regulated by fructose‐2,6‐bisphosphate; trypanothione as a substitute for glutathione; synthesis of fatty acids via a unique set of elongase enzymes; and a mitochondrial acetate:succinate coenzyme A transferase. B. saltans has lost the capacity to synthesize ubiquinone. Among genes that are present in B. saltans and lost in all trypanosomatids are those involved in the degradation of mureine, tryptophan and lysine. Novel acquisitions of trypanosomatids are components of pentose sugar metabolism, pteridine reductase and bromodomain‐factor proteins. In addition, only the subfamily Leishmaniinae has acquired a gene for catalase and the capacity to convert diaminopimelic acid to lysine.     

Synergistic regulation of immunoreceptor signaling by SLP-76-related adaptor Clnk and serine/threonine protein kinase HPK-1

Recently, the identification of Clnk, a third member of the SLP-76 family of adaptors expressed exclusively in cytokine-stimulated hemopoietic cells, has been reported by us and by others. Like SLP-76 and Blnk, Clnk was shown to act as a positive regulator of immunoreceptor signaling. Interestingly, however, it did not detectably associate with known binding partners of SLP-76, including Vav, Nck, and GADS. In contrast, it became complexed in activated T cells and myeloid cells with an as yet unknown tyrosine-phosphorylated polypeptide of approximately 92 kDa (p92). In order to understand better the function of Clnk, we sought to identify the Clnk-associated p92. Using a yeast two-hybrid screen and cotransfection experiments with Cos-1 cells, evidence was adduced that p92 is HPK-1, a serine/threonine-specific protein kinase expressed in hemopoietic cells. Further studies showed that Clnk and HPK-1 were also associated in hemopoietic cells and that their interaction was augmented by immunoreceptor stimulation. A much weaker association was detected between HPK-1 and SLP-76. Transient transfections in Jurkat T cells revealed that Clnk and HPK-1 cooperated to increase immunoreceptor-mediated activation of the interleukin 2 (IL-2) promoter. Moreover, the ability of Clnk to stimulate IL-2 promoter activity could be blocked by expression of a kinase-defective version of HPK-1. Lastly we found that in spite of the differential ability of Clnk and SLP-76 to bind cellular proteins, Clnk was apt at rescuing immunoreceptor signaling in a Jurkat T-cell variant lacking SLP-76. Taken together, these results show that Clnk physically and functionally interacts with HPK-1 in hemopoietic cells. Moreover, they suggest that Clnk is capable of functionally substituting for SLP-76 in immunoreceptor signaling, albeit by using a distinct set of intracellular effectors.     

Evaluating the Potential for the Environmentally Sustainable Control of Foot and Mouth Disease in Sub-Saharan Africa

Strategies to control transboundary diseases have in the past generated unintended negative consequences for both the environment and local human populations. Integrating perspectives from across disciplines, including livestock, veterinary and conservation sectors, is necessary for identifying disease control strategies that optimise environmental goods and services at the wildlife-livestock interface. Prompted by the recent development of a global strategy for the control and elimination of foot-and-mouth disease (FMD), this paper seeks insight into the consequences of, and rational options for potential FMD control measures in relation to environmental, conservation and human poverty considerations in Africa. We suggest a more environmentally nuanced process of FMD control that safe-guards the integrity of wild populations and the ecosystem dynamics on which human livelihoods depend while simultaneously improving socio-economic conditions of rural people. In particular, we outline five major issues that need to be considered: 1) improved understanding of the different FMD viral strains and how they circulate between domestic and wildlife populations; 2) an appreciation for the economic value of wildlife for many African countries whose presence might preclude the country from ever achieving an FMD-free status; 3) exploring ways in which livestock production can be improved without compromising wildlife such as implementing commodity-based trading schemes; 4) introducing a participatory approach involving local farmers and the national veterinary services in the control of FMD; and 5) finally the possibility that transfrontier conservation might offer new hope of integrating decision-making at the wildlife-livestock interface.     

An Economic Evaluation of TENS in Addition to Usual Primary Care Management for the Treatment of Tennis Elbow: Results from the TATE Randomized Controlled Trial

Background

The TATE trial was a multicentre pragmatic randomized controlled trial of supplementing primary care management (PCM)–consisting of a GP consultation followed by information and advice on exercises–with transcutaneous electrical nerve stimulation (TENS), to reduce pain intensity in patients with tennis elbow. This paper reports the health economic evaluation.

Methods and Findings

Adults with new diagnosis of tennis elbow were recruited from 38 general practices in the UK, and randomly allocated to PCM (n = 120) or PCM plus TENS (n = 121). Outcomes included reduction in pain intensity and quality-adjusted-life-years (QALYs) based on the EQ5D and SF6D. Two economic perspectives were evaluated: (i) healthcare–inclusive of NHS and private health costs for the tennis elbow; (ii) societal–healthcare costs plus productivity losses through work absenteeism. Mean outcome and cost differences between the groups were evaluated using a multiple imputed dataset as the base case evaluation, with uncertainty represented in cost-effectiveness planes and through probabilistic cost-effectiveness acceptability curves). Incremental healthcare cost was £33 (95%CI -40, 106) and societal cost £65 (95%CI -307, 176) for PCM plus TENS. Mean differences in outcome were: 0.11 (95%CI -0.13, 0.35) for change in pain (0–10 pain scale); -0.015 (95%CI -0.058, 0.029) for QALY_EQ5D; 0.007 (95%CI -0.022, 0.035) for QALY_SF6D (higher score differences denote greater benefit for PCM plus TENS). The ICER (incremental cost effectiveness ratio) for the main evaluation of mean difference in societal cost (£) relative to mean difference in pain outcome was -582 (95%CI -8666, 8113). However, incremental ICERs show differences in cost–effectiveness of additional TENS, according to the outcome being evaluated.

Conclusion

Our findings do not provide evidence for or against the cost-effectiveness of TENS as an adjunct to primary care management of tennis elbow.     

Structural insights and biomedical potential of IgNAR scaffolds from sharks

In addition to antibodies with the classical composition of heavy and light chains, the adaptive immune repertoire of sharks also includes a heavy-chain only isotype, where antigen binding is mediated exclusively by a small and highly stable domain, referred to as vNAR. In recent years, due to their high affinity and specificity combined with their small size, high physicochemical stability and low-cost of production, vNAR fragments have evolved as promising target-binding scaffolds that can be tailor-made for applications in medicine and biotechnology. This review highlights the structural features of vNAR molecules, addresses aspects of their generation using immunization or in vitro high throughput screening methods and provides examples of therapeutic, diagnostic and other biotechnological applications.