Thymic selection threshold defined by compartmentalization of Ras/MAPK signalling

A healthy individual can mount an immune response to exogenous pathogens while avoiding an autoimmune attack on normal tissues. The ability to distinguish between self and non-self is called 'immunological tolerance' and, for T lymphocytes, involves the generation of a diverse pool of functional T cells through positive selection and the removal of overtly self-reactive thymocytes by negative selection during T-cell ontogeny. To elucidate how thymocytes arrive at these cell fate decisions, here we have identified ligands that define an extremely narrow gap spanning the threshold that distinguishes positive from negative selection. We show that, at the selection threshold, a small increase in ligand affinity for the T-cell antigen receptor leads to a marked change in the activation and subcellular localization of Ras and mitogen-activated protein kinase (MAPK) signalling intermediates and the induction of negative selection. The ability to compartmentalize signalling molecules differentially in the cell endows the thymocyte with the ability to convert a small change in analogue input (affinity) into a digital output (positive versus negative selection) and provides the basis for establishing central tolerance.     

Structural diversity in binary nanoparticle superlattices

Assembly of small building blocks such as atoms, molecules and nanoparticles into macroscopic structures--that is, 'bottom up' assembly--is a theme that runs through chemistry, biology and material science. Bacteria, macromolecules and nanoparticles can self-assemble, generating ordered structures with a precision that challenges current lithographic techniques. The assembly of nanoparticles of two different materials into a binary nanoparticle superlattice (BNSL) can provide a general and inexpensive path to a large variety of materials (metamaterials) with precisely controlled chemical composition and tight placement of the components. Maximization of the nanoparticle packing density has been proposed as the driving force for BNSL formation, and only a few BNSL structures have been predicted to be thermodynamically stable. Recently, colloidal crystals with micrometre-scale lattice spacings have been grown from oppositely charged polymethyl methacrylate spheres. Here we demonstrate formation of more than 15 different BNSL structures, using combinations of semiconducting, metallic and magnetic nanoparticle building blocks. At least ten of these colloidal crystalline structures have not been reported previously. We demonstrate that electrical charges on sterically stabilized nanoparticles determine BNSL stoichiometry; additional contributions from entropic, van der Waals, steric and dipolar forces stabilize the variety of BNSL structures.     

Deconstruction of a neural circuit for hunger

Hunger is a complex behavioural state that elicits intense food seeking and consumption. These behaviours are rapidly recapitulated by activation of starvation-sensitive AGRP neurons, which present an entry point for reverse-engineering neural circuits for hunger. Here we mapped synaptic interactions of AGRP neurons with multiple cell populations in mice and probed the contribution of these distinct circuits to feeding behaviour using optogenetic and pharmacogenetic techniques. An inhibitory circuit with paraventricular hypothalamus (PVH) neurons substantially accounted for acute AGRP neuron-evoked eating, whereas two other prominent circuits were insufficient. Within the PVH, we found that AGRP neurons target and inhibit oxytocin neurons, a small population that is selectively lost in Prader-Willi syndrome, a condition involving insatiable hunger. By developing strategies for evaluating molecularly defined circuits, we show that AGRP neuron suppression of oxytocin neurons is critical for evoked feeding. These experiments reveal a new neural circuit that regulates hunger state and pathways associated with overeating disorders.     

Structural basis for iron piracy by pathogenic Neisseria

Neisseria are obligate human pathogens causing bacterial meningitis, septicaemia and gonorrhoea. Neisseria require iron for survival and can extract it directly from human transferrin for transport across the outer membrane. The transport system consists of TbpA, an integral outer membrane protein, and TbpB, a co-receptor attached to the cell surface; both proteins are potentially important vaccine and therapeutic targets. Two key questions driving Neisseria research are how human transferrin is specifically targeted, and how the bacteria liberate iron from transferrin at neutral pH. To address these questions, we solved crystal structures of the TbpA-transferrin complex and of the corresponding co-receptor TbpB. We characterized the TbpB-transferrin complex by small-angle X-ray scattering and the TbpA-TbpB-transferrin complex by electron microscopy. Our studies provide a rational basis for the specificity of TbpA for human transferrin, show how TbpA promotes iron release from transferrin, and elucidate how TbpB facilitates this process.     

Analysis of Plasmodium falciparum diversity in natural infections by deep sequencing

Malaria elimination strategies require surveillance of the parasite population for genetic changes that demand a public health response, such as new forms of drug resistance. Here we describe methods for the large-scale analysis of genetic variation in Plasmodium falciparum by deep sequencing of parasite DNA obtained from the blood of patients with malaria, either directly or after short-term culture. Analysis of 86,158 exonic single nucleotide polymorphisms that passed genotyping quality control in 227 samples from Africa, Asia and Oceania provides genome-wide estimates of allele frequency distribution, population structure and linkage disequilibrium. By comparing the genetic diversity of individual infections with that of the local parasite population, we derive a metric of within-host diversity that is related to the level of inbreeding in the population. An open-access web application has been established for the exploration of regional differences in allele frequency and of highly differentiated loci in the P.?falciparum genome.     

Warm tropical sea surface temperatures in the Late Cretaceous and Eocene epochs

Climate models with increased levels of carbon dioxide predict that global warming causes heating in the tropics, but investigations of ancient climates based on palaeodata have generally indicated cool tropical temperatures during supposed greenhouse episodes. For example, in the Late Cretaceous and Eocene epochs there is abundant geological evidence for warm, mostly ice-free poles, but tropical sea surface temperatures are generally estimated to be only 15-23 degrees C, based on oxygen isotope palaeothermometry of surface-dwelling planktonic foraminifer shells. Here we question the validity of most such data on the grounds of poor preservation and diagenetic alteration. We present new data from exceptionally well preserved foraminifer shells extracted from impermeable clay-rich sediments, which indicate that for the intervals studied, tropical sea surface temperatures were at least 28-32 degrees C. These warm temperatures are more in line with our understanding of the geographical distributions of temperature-sensitive fossil organisms and the results of climate models with increased CO2 levels.     

Mutations in a member of the Ras superfamily of small GTP-binding proteins causes Bardet-Biedl syndrome

RAB, ADP-ribosylation factors (ARFs) and ARF-like (ARL) proteins belong to the Ras superfamily of small GTP-binding proteins and are essential for various membrane-associated intracellular trafficking processes. None of the approximately 50 known members of this family are linked to human disease. Using a bioinformatic screen for ciliary genes in combination with mutational analyses, we identified ARL6 as the gene underlying Bardet-Biedl syndrome type 3, a multisystemic disorder characterized by obesity, blindness, polydactyly, renal abnormalities and cognitive impairment. We uncovered four different homozygous substitutions in ARL6 in four unrelated families affected with Bardet-Biedl syndrome, two of which disrupt a threonine residue important for GTP binding and function of several related small GTP-binding proteins. Analysis of the Caenorhabditis elegans ARL6 homolog indicates that it is specifically expressed in ciliated cells, and that, in addition to the postulated cytoplasmic functions of ARL proteins, it undergoes intraflagellar transport. These findings implicate a small GTP-binding protein in ciliary transport and the pathogenesis of a pleiotropic disorder.     

Loss of BBS proteins causes anosmia in humans and defects in olfactory cilia structure and function in the mouse

Defects in cilia are associated with several human disorders, including Kartagener syndrome, polycystic kidney disease, nephronophthisis and hydrocephalus. We proposed that the pleiotropic phenotype of Bardet-Biedl syndrome (BBS), which encompasses retinal degeneration, truncal obesity, renal and limb malformations and developmental delay, is due to dysfunction of basal bodies and cilia. Here we show that individuals with BBS have partial or complete anosmia. To test whether this phenotype is caused by ciliary defects of olfactory sensory neurons, we examined mice with deletions of Bbs1 or Bbs4. Loss of function of either BBS protein affected the olfactory, but not the respiratory, epithelium, causing severe reduction of the ciliated border, disorganization of the dendritic microtubule network and trapping of olfactory ciliary proteins in dendrites and cell bodies. Our data indicate that BBS proteins have a role in the microtubule organization of mammalian ciliated cells and that anosmia might be a useful determinant of other pleiotropic disorders with a suspected ciliary involvement.     

Lifespan: catch-up growth and obesity in male mice

Poor fetal growth is linked with long-term detrimental effects on health in adulthood. Here we investigate whether the lifespan of male mice is affected by their growth rate when they were suckling and find that limiting growth during that period not only increases longevity but also protects against the life-shortening effect of an obesity-inducing diet later on. By contrast, we find that lifespan is considerably shortened if the postnatal period of growth is accelerated to make up for reduced growth in utero, and that, in addition, these mice are susceptible to the adverse effects on longevity of an obesity-inducing diet after weaning.