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A search for a signal consistent with the type-III seesaw mechanism in events with three or more electrons or muons is presented. The data sample consists of proton-proton collisions at root s = 13 TeV collected by the CMS experiment at the LHC in 2016 and corresponds to an integrated luminosity of 35.9 fb(-1). Selection criteria based on the number of leptons and the invariant mass of oppositely charged lepton pairs are used to distinguish the signal from the standard model background. The observations are consistent with the expectations from standard model processes. The results are used to place limits on the production of heavy fermions of the type-III seesaw model as a function of the branching ratio to each lepton flavor. In the scenario of equal branching fractions to each lepton flavor, heavy fermions with masses below 840 GeV are excluded. This is the most sensitive probe to date of the type-III seesaw mechanism.

Mechanosensitive ion channels convert external mechanical stimuli into electrochemical signals for critical processes including touch sensation, balance, and cardiovascular regulation. The best understood mechanosensitive channel, MscL, opens a wide pore, which accounts for mechanosensitive gating due to in-plane area expansion. Eukaryotic Piezo channels have a narrow pore and therefore must capture mechanical forces to control gating in another way. We present a cryo-EM structure of mouse Piezo1 in a closed conformation at 3.7 angstrom-resolution. The channel is a triskelion with arms consisting of repeated arrays of 4-TM structural units surrounding a pore. Its shape deforms the membrane locally into a dome. We present a hypothesis in which the membrane deformation changes upon channel opening. Quantitatively, membrane tension will alter gating energetics in proportion to the change in projected area under the dome. This mechanism can account for highly sensitive mechanical gating in the setting of a narrow, cation-selective pore.

Calcium imaging with cellular resolution typically requires an animal to be tethered under a microscope, which substantially restricts the range of behaviors that can be studied. To expand the behavioral repertoire amenable to imaging, we have developed a tracking microscope that enables whole-brain calcium imaging with cellular resolution in freely swimming larval zebrafish. This microscope uses infrared imaging to track a target animal in a behavior arena. On the basis of the predicted trajectory of the animal, we applied optimal control theory to a motorized stage system to cancel brain motion in three dimensions. We combined this motion-cancellation system with differential illumination focal filtering, a variant of HiLo microscopy, which enabled us to image the brain of a freely swimming larval zebrafish for more than an hour. This work expands the repertoire of natural behaviors that can be studied with cellular-resolution calcium imaging to potentially include spatial navigation, social behavior, feeding and reward.

Background Staphylococcus aureus is a major human pathogen that can colonize healthy people mainly in the anterior nares. The aim of the present study was to evaluate S. aureus nasal colonization over time among Portuguese nursing students, including methicillin-resistant S. aureus (MRSA). Methods and findings In this longitudinal cohort study, we collected 280 nasal swabs from nursing students at 14 time points over four years of schooling (2012-2016). The isolates were characterized by pulsed-field gel electrophoresis (PFGE), spa typing, multilocus sequence typing (MLST), and SCCmec typing for MRSA. Among 47 students, 20 (43%) carried methicillin-susceptible S. aureus (MSSA) at admission, but none was colonized with MRSA. A total of 19 students (40%) became colonized after exposure during the nursing training, out of which five carried MRSA. Overall, 39 students (83%) had S. aureus detected at least once during the study period. Among the 97 MSSA isolates, most (65%) belonged to four clones: PFGE A-ST30 (21%), B-ST72 (20%), C-ST508 (13%), and D-ST398 (11%). Three of the five MRSA carriers were colonized with the predominant clone circulating in Portuguese hospitals (ST22-IVh) and two with ST3162-II. Colonization of nursing students was highly dynamic with continuous appearance of strains with distinct PFGE types in the same individual. Conclusions A considerable proportion of students became colonized by S. aureus, including MRSA, during the nursing education, evidencing this population represents an important reservoir of S. aureus. Therefore, education on infection control measures in nursing schools is of major importance.

S531 of Escherichia coli RNA polymerase (RNAP) beta subunit is a part of RNA binding domain in transcription complex. While highly conserved, S531 is not involved in interactions within the transcription complex as suggested by X-ray analysis. To understand the basis for S531 conservation we performed systematic mutagenesis of this residue. We find that the most of the mutations significantly decreased initiation-to-elongation transition by RNAP. Surprisingly, some changes enhanced the production of full-size transcripts by suppressing abortive loss of short RNAs. S531-R increased transcript retention by establishing a salt bridge with RNA, thereby explaining the R substitution at the equivalent position in extremophilic organisms, in which short RNAs retention is likely to be an issue. Generally, the substitutions had the same effect on bacterial doubling time when measured at 20 degrees. Raising growth temperature to 37 degrees ablated the positive influence of some mutations on the growth rate in contrast to their in vitro action, reflecting secondary effects of cellular environment on transcription and complex involvement of 531 locus in the cell biology. The properties of generated RNAP variants revealed an RNA/protein interaction network that is crucial for transcription, thereby explaining the details of initiation-to-elongation transition on atomic level.

Polycomb group proteins (PcG) are transcriptional repressors that control cell identity and development. In mammals, five different CBX proteins associate with the core Polycomb repressive complex 1 (PRC1). In mouse embryonic stem cells (ESCs), CBX6 and CBX7 are the most highly expressed CBX family members. CBX7 has been recently characterized, but little is known regarding the function of CBX6. Here, we show that CBX6 is essential for ESC identity. Its depletion destabilizes the pluripotency network and triggers differentiation. Mechanistically, we find that CBX6 is physically and functionally associated to both canonical PRC1 (cPRC1) and non-canonical PRC1 (ncPRC1) complexes. Notably, in contrast to CBX7, CBX6 is recruited to chromatin independently of H3K27me3. Taken together, our findings reveal that CBX6 is an essential component of ESC biology that contributes to the structural and functional complexity of the PRC1 complex.

Calcium phosphate-based mineralo-organic particles form spontaneously in the body and may represent precursors of ectopic calcification. We have shown earlier that these particles induce activation of caspase-1 and secretion of IL-1 beta by macrophages. However, whether the particles may produce other effects on immune cells is unclear. Here, we show that these particles induce the release of neutrophil extracellular traps (NETs) in a size-dependent manner by human neutrophils. Intracellular production of reactive oxygen species is required for particle-induced NET release by neutrophils. NETs contain the high-mobility group protein B1 (HMGB1), a DNA-binding protein capable of inducing secretion of TNF-alpha by a monocyte/macrophage cell line and primary macrophages. HMGB1 functions as a ligand of Toll-like receptors 2 and 4 on macrophages, leading to activation of the MyD88 pathway and TNF-alpha production. Furthermore, HMGB1 is critical to activate the particle-induced pro-inflammatory cascade in the peritoneum of mice. These results indicate that mineral particles promote pro-inflammatory responses by engaging neutrophils and macrophages via signaling of danger signals through NETs.

Intestinal intraepithelial lymphocytes (IELs) are located at the critical interface between the intestinal lumen, which is chronically exposed to food and microbes, and the core of the body. Using high-resolution microscopy techniques and intersectional genetic tools, we investigated the nature of IEL responses to luminal microbes. We observed that TCRgd IELs exhibit unique microbiota-dependent location and movement patterns in the epithelial compartment. This behavioral pattern quickly changes upon exposure to different enteric pathogens, resulting in increased interepithelial cell (EC) scanning, expression of antimicrobial genes, and glycolysis. Both dynamic and metabolic changes to gamma delta IEL depend on pathogen sensing by ECs. Direct modulation of glycolysis is sufficient to change gd IEL behavior and susceptibility to early pathogen invasion. Our results uncover a coordinated EC-IEL response to enteric infections that modulates lymphocyte energy utilization and dynamics and supports maintenance of the intestinal epithelial barrier.

Voltage-gated Na+ channels (Na-v) modulate neuronal excitability, but the roles of the various Na-v subtypes in specific neuronal functions such as synaptic transmission are unclear. We investigated expression of the three major brain Na-v subtypes (Na(v)1.1, Na(v)1.2, Na(v)1.6) in area CA1 and dentate gyrus of rat hippocampus. Using light and electron microscopy, we found labeling for all three Na-v subtypes on dendrites, dendritic spines, and axon terminals, but the proportion of pre- and post-synaptic labeling for each subtype varied within and between subregions of CA1 and dentate gyrus. In the central hilus (CH) of the dentate gyrus, Na(v)1.1 immunoreactivity was selectively expressed in presynaptic profiles, while Na(v)1.2 and Na(v)1.6 were expressed both pre- and post-synaptically. In contrast, in the stratum radiatum (SR) of CA1, Na(v)1.1, Na(v)1.2, and Na(v)1.6 were selectively expressed in postsynaptic profiles. We next compared differences in Na-v subtype expression between CH and SR axon terminals and between CH and SR dendrites and spines. Na(v)1.1 and Na(v)1.2 immunoreactivity was preferentially localized to CH axon terminals compared to SR, and in SR dendrites and spines compared to CH. No differences in Na(v)1.6 immunoreactivity were found between axon terminals of CH and SR or between dendrites and spines of CH and SR. All Na-v subtypes in both CH and SR were preferentially associated with asymmetric synapses rather than symmetric synapses. These findings indicate selective presynaptic and postsynaptic Na-v expression in glutamatergic synapses of CH and SR supporting neurotransmitter release and synaptic plasticity.