Sensory systems define an animal's capacity for perception and can evolve to promote survival in new environmental niches. We have uncovered a noncanonical mechanism for sweet taste perception that evolved in hummingbirds since their divergence from insectivorous swifts, their closest relatives. We observed the widespread absence in birds of an essential subunit (T1R2) of the only known vertebrate sweet receptor, raising questions about how specialized nectar feeders such as hummingbirds sense sugars. Receptor expression studies revealed that the ancestral umami receptor (the T1R1-T1R3 heterodimer) was repurposed in hummingbirds to function as a carbohydrate receptor. Furthermore, the molecular recognition properties of T1R1-T1R3 guided taste behavior in captive and wild hummingbirds. We propose that changing taste receptor function enabled hummingbirds to perceive and use nectar, facilitating the massive radiation of hummingbird species.
Structure determination of gold nanoparticles (AuNPs) is necessary for understanding their physical and chemical properties, and only one AuNP larger than 1 nm in diameter, an Au102NP, has been solved to atomic resolution. Whereas the Au102NP structure was determined by X-ray crystallography, other large AuNPs have proved refractory to this approach. Here we report the structure determination of an Au68NP at atomic resolution by aberration-corrected transmission electron microscopy (AC-TEM), performed with the use of a minimal electron dose, an approach that should prove applicable to metal NPs in general. The structure of the Au68NP was supported by small angle X-ray scattering (SAXS) and by comparison of observed infrared (IR) absorption spectra with calculations by density functional theory (DFT).
Patient-derived pluripotent stem cells (PSC) directed to various cell fates holds promise as source material for treating numerous disorders. The availability of precisely differentiated PSC-derived cells will dramatically impact blood component and hematopoietic stem cell therapies, and should facilitate treatment of diabetes, some forms of liver disease and neurologic disorders, retinal diseases, and possibly heart disease. Although an unlimited supply of specific cell types are needed, other barriers must be overcome. This review of the state of cell therapies highlights important challenges. Successful cell transplantation will require optimizing the best cell type and site for engraftment, overcoming limitations to cell migration and tissue integration, and occasionally needing to control immunologic reactivity. Collaboration among scientists, clinicians, and industry is critical for generating new stem cell-based therapies.
Flaviviruses, the human pathogens responsible for dengue fever, West Nile fever, tick-borne encephalitis and yellow fever, are endemic in tropical and temperate parts of the world. The flavivirus non-structural protein 1 (NS1) functions in genome replication as an intracellular dimer and in immune system evasion as a secreted hexamer. We report crystal structures for full-length, glycosylated NS1 from West Nile and dengue viruses. The NS1 hexamer in crystal structures is similar to a solution hexamer visualized by single-particle electron microscopy. Recombinant NS1 binds to lipid bilayers and remodels large liposomes into lipoprotein nanoparticles. The NS1 structures reveal distinct domains for membrane association of the dimer and interactions with the immune system, and are a basis for elucidating the molecular mechanism of NS1 function.
Decapentaplegic (Dpp), a Drosophila morphogen signaling protein, transfers directly at synapes made at sites of contact between cells that produce Dpp and cytonemes that extend from recipient cells. The Dpp that cytonemes receive moves together with activated receptors toward the recipient cell body in motile puncta. Genetic loss-of-function conditions for diaphanous, shibire, neuroglian and capricious perturbed cytonemes by reducing their number or only the synapses they make with cells they target; and reduced cytoneme-mediated transport of Dpp and Dpp signaling. These experiments provide direct evidence that cells use cytonemes to exchange signaling proteins, that cytoneme-based exchange is essential for signaling and normal development, and that morphogen distribution and signaling can be contact-dependent, requiring cytoneme synapses.
Drosophila cytonemes; Dpp signaling; morphogen; synapse
Although fluorescence microscopy provides a crucial window into the physiology of living specimens, many biological processes are too fragile, too small, or occur too rapidly to see clearly with existing tools. We crafted ultra-thin light sheets from two-dimensional optical lattices that allowed us to image three-dimensional (3D) dynamics for hundreds of volumes, often at sub-second intervals, at the diffraction limit and beyond. We applied this to systems spanning four orders of magnitude in space and time, including the diffusion of single transcription factor molecules in stem cell spheroids, the dynamic instability of mitotic microtubules, the immunological synapse, neutrophil motility in a 3D matrix, and embryogenesis in Caenorhabditis elegans and Drosophila melanogaster. The results provide a visceral reminder of the beauty and complexity of living systems.
Clathrin-mediated endocytosis (CME) is vital for the internalization of most cell-surface proteins. In CME, plasma membrane-binding clathrin adaptors recruit and polymerize clathrin to form clathrin-coated ‘pits’ into which cargo is sorted. AP2 is the most abundant adaptor, and is pivotal to CME. By determining a new structure of AP2 that includes the clathrin-binding β2-hinge and developing an AP2-dependent budding assay, we reveal the existence of an autoinhibitory mechanism that prevents clathrin recruitment by cytosolic AP2. A large-scale conformational change driven by the plasma membrane phosphoinositide PtdIns(4,5)P2 and cargo relieves this autoinhibition, so triggering clathrin recruitment and hence clathrin-coated bud formation. This molecular switching mechanism constitutes an unsuspected layer of regulation that couples AP2’s membrane recruitment to its key functions of cargo and clathrin binding.
Cellular circuits sense the environment, process signals, and compute decisions using networks of interacting proteins. To model such a system, the abundance of each activated protein species can be described as a stochastic function of the abundance of other proteins. High-dimensional single-cell technologies, like mass cytometry, offer an opportunity to characterize signaling circuit-wide. However, the challenge of developing and applying computational approaches to interpret such complex data remains. Here, we developed computational methods, based on established statistical concepts, to characterize signaling network relationships by quantifying the strengths of network edges and deriving signaling response functions. In comparing signaling between naïve and antigen-exposed CD4+ T-lymphocytes, we find that although these two cell subtypes had similarly-wired networks, naïve cells transmitted more information along a key signaling cascade than did antigen-exposed cells. We validated our characterization on mice lacking the extracellular-regulated MAP kinase (ERK2), which showed stronger influence of pERK on pS6 (phosphorylated-ribosomal protein S6), in naïve cells compared to antigen-exposed cells, as predicted. We demonstrate that by using cell-to-cell variation inherent in single cell data, we can algorithmically derive response functions underlying molecular circuits and drive the understanding of how cells process signals.
Typical therapies try to reverse pathogenic mechanisms. Here, we describe treatment effects by enhancing depression-causing mechanisms in ventral tegmental area (VTA) dopamine (DA) neurons. In a social defeat stress model of depression, depressed (susceptible) mice display hyperactivity of VTA DA neurons, caused by an up-regulated hyperpolarization-activated current (Ih). Mice resilient to social defeat stress, however, exhibit stable normal firing of these neurons. Unexpectedly, resilient mice had an even larger Ih, which was observed in parallel with increased potassium (K+) channel currents. Experimentally enhancing the firing-increasing Ih or optogenetically increasing the hyperactivity of VTA DA neurons in susceptible mice, completely reversed depression-related behaviors, an antidepressant effect achieved through resilience-like, projection-specific homeostatic plasticity. These results indicate a potential therapeutic path of promoting natural resilience for depression treatment.
Defensins are antimicrobial peptides that contribute broadly to innate immunity, including protection of mucosal tissues. Human α-defensin (HD)6 is highly expressed by secretory Paneth cells of the small intestine. However, in contrast to the other defensins, it lacks appreciable bactericidal activity. Nevertheless, we report here that HD6 affords protection against invasion by enteric bacterial pathogens in vitro and in vivo. After stochastic binding to bacterial surface proteins, HD6 undergoes ordered self-assembly to form fibrils and nanonets that surround and entangle bacteria. This self-assembly mechanism occurs in vivo, requires histidine-27, and is consistent with X-ray crystallography data. These findings support a key role for HD6 in protecting the small intestine against invasion by diverse enteric pathogens, and may explain the conservation of HD6 throughout Hominidae evolution.
In 11 studies, we found that participants typically did not enjoy spending 6 to 15 minutes in a room by themselves with nothing to do but think, that they enjoyed doing mundane external activities much more, and that many preferred to administer electric shocks to themselves instead of being left alone with their thoughts. Most people seem to prefer to be doing something rather than nothing, even if that something is negative.
“The mind is its own place, and in it self/Can make a Heav'n of Hell, a Hell of Heav'n.”– John Milton, Paradise Lost
The formation of body segments (somites) in vertebrate embryos is accompanied by molecular oscillations (segmentation clock). Interaction of this oscillator with a wave traveling along the body axis (the clock-and-wavefront model) is generally believed to control somite number, size, and axial identity. Here we show that a clock-and-wavefront mechanism is unnecessary for somite formation. Non-somite mesoderm treated with Noggin generates many somites that form simultaneously, without cyclic expression of Notch-pathway genes, yet have normal size, shape, and fate. These somites have axial identity: The Hox code is fixed independently of somite fate. However, these somites are not subdivided into rostral and caudal halves, which is necessary for neural segmentation. We propose that somites are self-organizing structures whose size and shape is controlled by local cell-cell interactions.
Articular cartilage was predicted to be one of the first tissues that could successfully be regenerated, but this proved not to be the case. In contrast, bone but also vasculature and cardiac tissues have seen numerous successful reparative approaches, despite consisting of multiple cell and tissue types and thus possessing more complex design requirements. Here, we use bone regeneration successes to highlight cartilage regeneration challenges, namely selecting appropriate cell sources and scaffolds, creating biomechanically suitable tissues, and integrating to native tissue. Also discussed are technologies addressing hurdles of engineering a tissue possessing mechanical properties unmatched in man-made materials and functioning in environments unfavorable to neotissue growth.
Small molecules inhibit a mutant enzyme confined to tumors, supporting
therapeutic approaches that can reprogram metabolism in cancer.
The toxicity of ionizing radiation is associated with massive apoptosis in radiosensitive organs. Here, we investigate whether a drug that activates a signaling mechanism used by tumor cells to suppress apoptosis can protect healthy cells from the harmful effects of radiation. We studied CBLB502, a polypeptide drug derived from Salmonella flagellin that binds to Toll-like receptor 5 (TLR5) and activates nuclear factor–κB signaling. A single injection of CBLB502 before lethal total-body irradiation protected mice from both gastrointestinal and hematopoietic acute radiation syndromes and resulted in improved survival. CBLB502 injected after irradiation also enhanced survival, but at lower radiation doses. It is noteworthy that the drug did not decrease tumor radiosensitivity in mouse models. CBLB502 also showed radioprotective activity in lethally irradiated rhesus monkeys. Thus, TLR5 agonists could potentially improve the therapeutic index of cancer radiotherapy and serve as biological protectants in radiation emergencies.
Type-I interferon (IFN) protects against viruses yet it also has a poorly understood suppressive influence on inflammation. Here we report that activated mouse macrophages lacking the IFN-stimulated gene, cholesterol 25-hydroxylase (Ch25h), and that are unable to produce the oxysterol 25-hydroxycholesterol (25-HC) overproduce inflammatory interleukin 1 (IL-1) family cytokines. 25-HC acts by antagonizing sterol response element-binding protein (SREBP) processing to reduce Il1b transcription and to broadly repress IL1-activating inflammasomes. In accord with these dual actions of 25-HC, Ch25h-deficient mice exhibit increased sensitivity to septic shock, exacerbated experimental autoimmune encephalomyelitis, and a stronger ability to repress bacterial growth. These findings identify an oxysterol, 25-HC, as a critical mediator in the negative-feedback pathway of IFN signaling on IL1-family cytokine production and inflammasome activity.
We report the discovery and crystal structure of a human mycoplasma protein, Protein M, which binds with high affinity to antibodies, predominantly through attachment to the variable region of the κ and λ light chains. Protein M broadly blocks antibody-antigen union and its mechanism of inhibition is of considerable interest because, as a diversity system, the binding mode of each antibody is different. Protein M thus appears to function by a mechanism that is independent of the sequences of members of the extensive antibody repertoire. By anchoring to conserved regions of the antibody light chains, Protein M is in a position to extend its large C-terminal domain over the antibody combining site and block entrance to macromolecular antigens.
Neuronal chloride concentration [Cl−]i is an important determinant of GABAA receptor (GABAAR)-mediated inhibition and cytoplasmic volume regulation. Equilibrative cation-chloride cotransporters (CCC) move Cl− across the membrane, but accumulating evidence suggests factors other than the bulk concentrations of transported ions determine [Cl−]i. Measurement of [Cl−]i in murine brain slice preparations expressing the transgenic fluorophore Clomeleon demonstrated that cytoplasmic impermeant anions ([A]i) and polyanionic extracellular matrix glycoproteins ([A]o) constrain the local [Cl−]. CCC inhibition had modest effects on [Cl−]i and neuronal volume, but substantial changes were produced by alterations of the balance between [A]i and [A]o. Therefore, CCC are important elements of Cl− homeostasis, but local impermeant anions determine the homeostatic set-point for [Cl−], and hence, neuronal volume and the polarity of local GABAAR signaling.
Rare genetic variants contribute to complex disease risk; however, the abundance of rare variants in human populations remains unknown. We explored this spectrum of variation by sequencing 202 genes encoding drug targets in 14,002 individuals. We find rare variants are abundant (one every 17 bases) and geographically localized, such that even with large sample sizes, rare variant catalogs will be largely incomplete. We used the observed patterns of variation to estimate population growth parameters, the proportion of variants in a given frequency class that are putatively deleterious, and mutation rates for each gene. Overall we conclude that, due to rapid population growth and weak purifying selection, human populations harbor an abundance of rare variants, many of which are deleterious and have relevance to understanding disease risk.
The late phase of long-term potentiation (LTP) at glutamatergic synapses, which
is thought to underlie long-lasting memory, requires gene transcription in the nucleus.
However, the mechanism by which signaling initiated at synapses is transmitted into the
nucleus to induce transcription has remained elusive. Here, we found that induction of LTP
in only a few dendritic spines was sufficient to activate extracellular signal-related
kinase (ERK) in the nucleus and regulate downstream transcription factors. Signaling from
individual spines was integrated over a wide range of time (>30 min) and space
(>80 μm). Spatially dispersed inputs over multiple branches activated
nuclear ERK much more efficiently than clustered inputs over one branch. Thus, biochemical
signals from individual dendritic spines exert profound effects on nuclear signaling.
Adipocytes have been suggested to be immunologically active, but their role in host defense is unclear. We observed rapid proliferation of preadipocytes and expansion of the dermal fat layer after infection of the skin by Staphylococcus aureus. Impaired adipogenesis resulted in increased infection as seen in Zfp423nur12 mice or in mice given inhibitors of peroxisome proliferator–activated receptor γ. This host defense function was mediated through the production of cathelicidin antimicrobial peptide from adipocytes because cathelicidin expression was decreased by inhibition of adipogenesis, and adipocytes from Camp−/− mice lost the capacity to inhibit bacterial growth. Together, these findings show that the production of an antimicrobial peptide by adipocytes is an important element for protection against S. aureus infection of the skin.
Chromosome fragmentation is a hallmark of apoptosis, conserved in diverse organisms. In mammals, caspases activate apoptotic chromosome fragmentation by cleaving and inactivating an apoptotic nuclease inhibitor. We report that inactivation of the C. elegans dcr-1 gene, which encodes the Dicer ribonuclease important for processing of small RNAs, compromises apoptosis and blocks apoptotic chromosome fragmentation. DCR-1 was cleaved by the CED-3 caspase to generate a C-terminal fragment with deoxyribonuclease activity, which produced 3’ hydroxyl DNA breaks on chromosomes and promoted apoptosis. Thus caspase-mediated activation of apoptotic DNA degradation is conserved. DCR-1 functions in fragmenting chromosomal DNA during apoptosis, in addition to processing of small RNAs, and undergoes a protease-mediated conversion from a ribonuclease to a deoxyribonuclease.
How the immune system adapts to malnutrition to sustain immunity at barrier surfaces such as the intestine remains unclear. Vitamin A deficiency is one of the most common micronutrient deficiencies, associated with profound defects in adaptive immunity. Here, we found that type 3 innate lymphoid cells (ILC3) are severely diminished under vitamin A deficient settings resulting in compromised immunity to acute bacterial infection. However, deprivation of vitamin A paradoxically resulted in dramatic expansion of IL-13 producing ILC2 and resistance to nematode infection revealing ILC as primary sensors of dietary stress. Further, these data propose that during malnutrition, a switch to innate type 2 immunity may represent a powerful adaptation of the immune system to promote host survival in the face of ongoing barrier challenges.