Design and use of conditional MHC class I ligands

Major histocompatibility complex (MHC) class I molecules associate with a variety of peptide ligands during biosynthesis and present these ligands on the cell surface for recognition by cytotoxic T cells. We have designed conditional MHC ligands that form stable complexes with MHC molecules but degrade on command, by exposure to a defined photostimulus. 'Empty MHC molecules' generated in this manner can be loaded with arrays of peptide ligands to determine MHC binding properties and to monitor antigen-specific T-cell responses in a high-throughput manner. We document the value of this approach by identifying cytotoxic T-cell epitopes within the H5N1 influenza A/Vietnam/1194/04 genome.     

Retinoic acid reduces the cytotoxicity of cyclopentenyl cytosine in neuroblastoma cells

To know the function of the Ca2+ channel containing alpha(1)2.3 (alpha1E) subunit (Ca(v)2.3 channel) in spermatozoa, we analyzed Ca2+ transients and sperm motility using a mouse strain lacking Ca(v)2.3 channel. The averaged rising rates of Ca2+ transients induced by alpha-D-mannose-bovine serum albumin in the head region of Ca(v)2.3-/- sperm were significantly lower than those of Ca(v)2.3+/+ sperm. A computer-assisted sperm motility assay revealed that straight-line velocity and linearity were greater in Ca(v)2.3-/- sperm than those in Ca(v)2.3+/+ sperm. These results suggest that the Ca(v)2.3 channel plays some roles in Ca2+ transients and the control of flagellar movement.     

Microwave-assisted diastereoselective two-step three-component synthesis for rapid access to drug-like libraries of substituted 3-amino-β-lactams

Large, diverse compound libraries are an essential requisite in target-based drug development. In this work, a robust microwave-assisted synthesis for the diastereoselective generation of 3-saccharinyl-trans-β-lactams is reported. The method is optimised for combinatorial library synthesis in which decoration of the scaffold is varied on both the β-lactam and the saccharine moiety. Within the European Lead Factory (ELF) consortium, a library of 263 compounds was efficiently produced using the developed methodology.     

Target Specificity of the E3 Ligase LUBAC for Ubiquitin and NEMO Relies on Different Minimal Requirements

The ubiquitination of NEMO with linear ubiquitin chains by the E3-ligase LUBAC is important for the activation of the canonical NF-κB pathway. NEMO ubiquitination requires a dual target specificity of LUBAC, priming on a lysine on NEMO and chain elongation on the N terminus of the priming ubiquitin. Here we explore the minimal requirements for these specificities. Effective linear chain formation requires a precise positioning of the ubiquitin N-terminal amine in a negatively charged environment on the top of ubiquitin. Whereas the RBR-LDD region on HOIP is sufficient for targeting the ubiquitin N terminus, the priming lysine modification on NEMO requires catalysis by the RBR domain of HOIL-1L as well as the catalytic machinery of the RBR-LDD domains of HOIP. Consequently, target specificity toward NEMO is determined by multiple LUBAC components, whereas linear ubiquitin chain elongation is realized by a specific interplay between HOIP and ubiquitin.     

Ant Community Development on Rehabilitated Ash Dams in the South African Highveld

Ant communities have been widely used as indicators of minesite rehabilitation in Australia and are beginning to play a similar role in other parts of the world. Here we examine ant communities on rehabilitated ash dams associated with a coal‐fired power station on the highveld of South Africa, to improve our understanding of ecosystem development on these substrates. Ants were sampled using pitfall traps at 11 ash‐dam sites, ranging from unrehabilitated to 9‐year‐old rehabilitated sites, as well as two adjacent natural grassland sites. Sampling was conducted on 12 occasions from March 1997 to January 1999. Forty‐nine ant species from 19 genera were recorded during the study. Site species richness was positively correlated with rehabilitation age, ranging from 10 to 25 at ash‐dam sites, compared with 28 and 34 at the two natural grassland sites. There was a humped relationship between total ant abundance and rehabilitation age, with abundance peaking after 5–7 years at levels far higher than those at natural sites. Ordination analysis showed clear separation between ash‐dam and natural sites along the first axis. The unrehabilitated ash‐dam site was also separated from rehabilitated sites along the first axis. Sites of different rehabilitation age were separated along the second axis. Individual ant species showed clear successional patterns across the rehabilitation gradient. Although there was a clear successional trend for the development of ant communities on rehabilitated ash dams, this trend was not toward natural grassland. The lack of convergence toward ant communities of natural grasslands reflects the markedly different substrate and plant composition on ash dams and supports the widely held view that restoration of natural grassland communities is not a realistic goal of ash‐dam rehabilitation. However, the development of species‐rich ant communities, containing at least some late‐successional species, indicates the potential for rehabilitated ash dams to support diverse and complex ecosystems.     

Technologies for MHC class I immunoproteomics

T cell epitopes are peptides, for instance derived from foreign, mutated or overexpressed proteins, that are displayed by MHC molecules on the cell surface and that are recognized by T lymphocytes. Knowledge of the identity of epitopes displayed by MHC molecules is of high value for diagnostic purposes and for the development of prophylactic and therapeutic immunotherapy regimens. Here we review key techniques in MHC class I epitope definition and we discuss recent developments in epitope discovery and their implications. Developments in epitope discovery strategies should ultimately lead to the definition of the MHC-associated peptidome.