Multimodal fast optical interrogation of neural circuitry

Our understanding of the cellular implementation of systems-level neural processes like action, thought and emotion has been limited by the availability of tools to interrogate specific classes of neural cells within intact, living brain tissue. Here we identify and develop an archaeal light-driven chloride pump (NpHR) from Natronomonas pharaonis for temporally precise optical inhibition of neural activity. NpHR allows either knockout of single action potentials, or sustained blockade of spiking. NpHR is compatible with ChR2, the previous optical excitation technology we have described, in that the two opposing probes operate at similar light powers but with well-separated action spectra. NpHR, like ChR2, functions in mammals without exogenous cofactors, and the two probes can be integrated with calcium imaging in mammalian brain tissue for bidirectional optical modulation and readout of neural activity. Likewise, NpHR and ChR2 can be targeted together to Caenorhabditis elegans muscle and cholinergic motor neurons to control locomotion bidirectionally. NpHR and ChR2 form a complete system for multimodal, high-speed, genetically targeted, all-optical interrogation of living neural circuits.     

Co-occurrence of three Aristolochia-feeding Papilionids (Archon apollinus,Zerynthia polyxena and Zerynthia cerisy) in Greek Thrace

Comparative studies of co-occurring species using overlapping resources may help in understanding the mechanisms supporting biotic diversity in species-rich regions, such as the Mediterranean region of Europe. Three Papilionidae butterflies, Archon apollinus, Zerynthia cerisy and Zerynthia polyxena, develop on Aristolochia plants and co-occur in Greek Thrace. We used mark–recapture to describe adult demography and dispersal, and searched for eggs and larvae to assess host plants and microhabitat preferences. Adult flight timing followed a sequence from earliest A. apollinus, through Z. polyxena to late Z. cerisy; this was more prominent in 2010 (warm early spring) than in 2011 (cold delayed spring). Population densities were highest for A. apollinus and lowest for Z. cerisy, whereas dispersal ability followed a reverse pattern. Adults of all three species crossed distances > 3 km and used all habitat types present. Four Aristolochia host plants were used at the study locality: small Aristolochia pallida, intermediate Aristolochia rotunda and Aristolochia hirta, and bulky, late-sprouting Aristolochia clematitis. Both A. apollinus and Z. polyxena used all four Aristolochia species, the former preferring Aristolochia rotunda and Aristolochia hirta, the latter Aristolochia rotunda and Aristolochia pallida. Zerynthia cerisy did not use the early-growing Aristolochia pallida while frequently using the late-growing Aristolochia clematitis. Further parameters affecting oviposition were biotope and canopy closure: early A. apollinus tolerated shady sites but late Z. cerisy avoided them. The simultaneous use of several host plants differing in phenology and habitat requirements, combined with rather high dispersal ability, arguably buffers the butterflies’ population dynamics against yearly variation in weather, while allowing efficient occupation of the diverse Mediterranean landscapes. The regional habitat diversity, created during millennia of human activity, is currently threatened by land abandonment, which may diminish the resource base for the studied butterflies.     

Evidence of G-protein-coupled receptor and substrate transporter heteromerization at a single molecule level

G-protein-coupled receptors (GPCRs) can constitute complexes with non-GPCR integral membrane proteins, while such interaction has not been demonstrated at a single molecule level so far. We here investigated the potential interaction between the thyrotropin receptor (TSHR) and the monocarboxylate transporter 8 (MCT8), a member of the major facilitator superfamily (MFS), using fluorescence cross-correlation spectroscopy (FCCS). Both the proteins are expressed endogenously on the basolateral plasma membrane of the thyrocytes and are involved in stimulation of thyroid hormone production and release. Indeed, we demonstrate strong interaction between both the proteins which causes a suppressed activation of G_q/11 by TSH-stimulated TSHR. Thus, we provide not only evidence for a novel interaction between the TSHR and MCT8, but could also prove this interaction on a single molecule level. Moreover, this interaction forces biased signaling at the TSHR. These results are of general interest for both the GPCR and the MFS research fields.     

<Emphasis Type="Italic">Yersinia pseudotuberculosis</Emphasis> supports Th17 differentiation and limits de novo regulatory T cell induction by directly interfering with T cell receptor signaling

Adaptive immunity critically contributes to control acute infection with enteropathogenic Yersinia pseudotuberculosis; however, the role of CD4⁺ T cell subsets in establishing infection and allowing pathogen persistence remains elusive. Here, we assessed the modulatory capacity of Y. pseudotuberculosis on CD4⁺ T cell differentiation. Using in vivo assays, we report that infection with Y. pseudotuberculosis resulted in enhanced priming of IL-17-producing T cells (Th17 cells), whereas induction of Foxp3⁺ regulatory T cells (Tregs) was severely disrupted in gut-draining mesenteric lymph nodes (mLNs), in line with altered frequencies of tolerogenic and proinflammatory dendritic cell (DC) subsets within mLNs. Additionally, by using a DC-free in vitro system, we could demonstrate that Y. pseudotuberculosis can directly modulate T cell receptor (TCR) downstream signaling within naïve CD4⁺ T cells and Tregs via injection of effector molecules through the type III secretion system, thereby affecting their functional properties. Importantly, modulation of naïve CD4⁺ T cells by Y. pseudotuberculosis resulted in an enhanced Th17 differentiation and decreased induction of Foxp3⁺ Tregs in vitro. These findings shed light to the adjustment of the Th17-Treg axis in response to acute Y. pseudotuberculosis infection and highlight the direct modulation of CD4⁺ T cell subsets by altering their TCR downstream signaling.     

Protein folding in membranes

Separation of cells and organelles by bilayer membranes is a fundamental principle of life. Cellular membranes contain a baffling variety of proteins, which fulfil vital functions as receptors and signal transducers, channels and transporters, motors and anchors. The vast majority of membrane-bound proteins contain bundles of α-helical transmembrane domains. Understanding how these proteins adopt their native, biologically active structures in the complex milieu of a membrane is therefore a major challenge in today’s life sciences. Here, we review recent progress in the folding, unfolding and refolding of α-helical membrane proteins and compare the molecular interactions that stabilise proteins in lipid bilayers. We also provide a critical discussion of a detergent denaturation assay that is increasingly used to determine membrane-protein stability but is not devoid of conceptual difficulties.     

A spontaneous chicken chimaera

Summary Among hybrids of the inbred lines, a chicken was found to be chimaeric in red blood cells and skin cells and to produce 3 types of sperm cells. This bird could have originated either from the fusion of 2 blastoderms or from the fertilization and fusion of the oocyte and the 2nd polar body.The authors wish to thank Professor Morten Simonsen. Institute for Experimental Immunology, University of Copenhagen, Copenhagen, and Dr Vincenzo Miggiano, Basel Institute for Immunology, Basel for stimulating and critical discussion, and Professor Milan Haek, Institute of Molecular Genetics, Czechoslovak Academy of Sciences, Prague, for his interest and advice.     

中国哲学概念里的"法"--从早期哲学经典到法家对"法"的诠释比较

文章旨在简要分析“法”字在早期中国哲学典籍及法家著述中的用法，试图证明在后来的法家著作中“法”字在法律、正义、惩罚等意义内涵上并没有改变早期哲学经典的诠释．