The rockefeller university

Alternative polyadenylation (APA) regulates mRNA translation, stability, and protein localization. However, it is unclear to what extent APA regulates these processes uniquely in specific cell types. Using a new technique, cTag-PAPERCLIP, we discovered significant differences in APA between the principal types of mouse cerebellar neurons, the Purkinje and granule cells, as well as between proliferating and differentiated granule cells. Transcripts that differed in APA in these comparisons were enriched in key neuronal functions and many differed in coding sequence in addition to 3'UTR length. We characterize Memo1, a transcript that shifted from expressing a short 3'UTR isoform to a longer one during granule cell differentiation. We show that Memo1 regulates granule cell precursor proliferation and that its long 3'UTR isoform is targeted by miR-124, contributing to its downregulation during development. Our findings provide insight into roles for APA in specific cell types and establish a platform for further functional studies.

Using 5' rapid amplification of cDNA ends, Illumina MiSeq, and basic flow cytometry, we systematically analyzed the expressed B cell receptor (BCR) repertoire in 14 healthy adult PBMCs, 5 HIV-1+adult PBMCs, 5 cord blood samples, and 3 HIS-CD4/B mice, examining the full-length variable region of mu, gamma, alpha, kappa, and lambda chains for V-gene usage, somatic hypermutation (SHM), and CDR3 length. Adding to the known repertoire of healthy adults, Illumina MiSeq consistently detected small fractions of reads with high mutation frequencies including hypermutated mu reads, and reads with long CDR3s. Additionally, the less studied IgA repertoire displayed similar characteristics to that of IgG. Compared to healthy adults, the five HIV-1 chronically infected adults displayed elevated mutation frequencies for all mu, gamma, alpha, kappa, and lambda chains examined and slightly longer CDR3 lengths for gamma, alpha, and lambda. To evaluate the reconstituted human BCR sequences in a humanized mouse model, we analyzed cord blood and HIS-CD4/B mice, which all lacked the typical SHM seen in the adult reference. Furthermore, MiSeq revealed identical unmutated IgM sequences derived from separate cell aliquots, thus for the first time demonstrating rare clonal members of unmutated IgM B cells by sequencing.

Events with no charged particles produced between the two leading jets are studied in proton-proton collisions at root s = 7 TeV. The jets were required to have transverse momentum p(T)(jet) > 40 GeV and pseudorapidity 1.5 < vertical bar eta(jet)vertical bar < 4.7, and to have values of eta(jet) with opposite signs. The data used for this study were collected with the CMSdetector during low-luminosity running at the LHC, and correspond to an integrated luminosity of 8 pb(-1). Events with no charged particles with p(T) > 0.2GeV in the interval -1 < eta < 1 between the jets are observed in excess of calculations that assume no color-singlet exchange. The fraction of events with such a rapidity gap, amounting to 0.5-1% of the selected dijet sample, is measured as a function of the pT of the second-leading jet and of the rapidity separation between the jets. The data are compared to previous measurements at the Tevatron, and to perturbative quantum chromodynamics calculations based on the Balitsky-Fadin-Kuraev-Lipatov evolution equations, including different models of the non-perturbative gap survival probability.

There is great interest in passive transfer of broadly neutralizing antibodies (bnAbs) and engineered bispecific antibodies (Abs) for prevention of HIV-1 infections due to their in vitro neutralization breadth and potency against global isolates and long in vivo half-lives. We compared the potential of eight bnAbs and two bispecific Abs currently under clinical development, and their 2 Ab combinations, to prevent infection by dominant HIV-1 subtypes in sub-Saharan Africa. Using in vitro neutralization data for Abs against 25 subtype A, 100 C, and 20 D pseudoviruses, we modeled neutralization by single Abs and 2 Ab combinations assuming realistic target concentrations of 10 mu g/ml total for bnAbs and combinations, and 5 mu g/ml for bispecifics. We used IC80 breadth-potency, completeness of neutralization, and simultaneous coverage by both Abs in the combination as metrics to characterize prevention potential. Additionally, we predicted in vivo protection by Abs and combinations by modeling protection as a function of in vitro neutralization based on data from a macaque simian-human immunodeficiency virus (SHIV) challenge study. Our model suggests that nearly complete neutralization of a given virus is needed for in vivo protection (similar to 98% neutralization for 50% relative protection). Using the above metrics, we found that bnAb combinations should outperform single bnAbs, as expected; however, different combinations are optimal for different subtypes. Remarkably, a single bispecific 10E8-iMAb, which targets HIV Env and host-cell CD4, outperformed all combinations of two conventional bnAbs, with 95-97% predicted relative protection across subtypes. Combinations that included 10E8-iMAb substantially improved protection over use of 10E8-iMAb alone. Our results highlight the promise of 10E8-iMAb and its combinations to prevent HIV-1 infections in sub-Saharan Africa.

A search for the production of Higgs boson pairs in proton-proton collisions at a centre-of-mass energy of 13TeVis presented, using a data sample corresponding to an integrated luminosity of 35.9fb(-1) collected with the CMS detector at the LHC. Events with one Higgs boson decaying into two bottom quarks and the other decaying into two tau leptons are explored to investigate both resonant and nonresonant production mechanisms. The data are found to be consistent, within uncertainties, with the standard model background predictions. For resonant production, upper limits at the 95% confidence level are set on the production cross section for Higgs boson pairs as a function of the hypothesized resonance mass and are interpreted in the context of the minimal supersymmetric standard model. For nonresonant production, upper limits on the production cross section constrain the parameter space for anomalous Higgs boson couplings. The observed (expected) upper limit at 95% confidence level corresponds to about 30(25) times the prediction of the standard model. (C) 2018 The Author. Published by Elsevier B.V.

Nuclear pore complexes play central roles as gatekeepers of RNA and protein transport between the cytoplasm and nucleoplasm. However, their large size and dynamic nature have impeded a full structural and functional elucidation. Here we determined the structure of the entire 552-protein nuclear pore complex of the yeast Saccharomyces cerevisiae at sub-nanometre precision by satisfying a wide range of data relating to the molecular arrangement of its constituents. The nuclear pore complex incorporates sturdy diagonal columns and connector cables attached to these columns, imbuing the structure with strength and flexibility. These cables also tie together all other elements of the nuclear pore complex, including membrane-interacting regions, outer rings and RNA-processing platforms. Inwardly directed anchors create a high density of transport factor-docking Phe-Gly repeats in the central channel, organized into distinct functional units. This integrative structure enables us to rationalize the architecture, transport mechanism and evolutionary origins of the nuclear pore complex.

Tumor metabolism is reorganized to support proliferation in the face of growth-related stress. Unlike the widespread profiling of changes to metabolic enzyme levels in cancer, comparatively less attention has been paid to the substrates/products of enzyme-catalyzed reactions, small-molecule metabolites. We developed an informatic pipeline to concurrently analyze metabolomics data from over 900 tissue samples spanning seven cancer types, revealing extensive heterogeneity in metabolic changes relative to normal tissue across cancers of different tissues of origin. Despite this heterogeneity, a number of metabolites were recurrently differentially abundant across many cancers, such as lactate and acyl-carnitine species. Through joint analysis of metabolomic data alongside clinical features of patient samples, we also identified a small number of metabolites, including several polyamines and kynurenine, which were associated with aggressive tumors across several tumor types. Our findings offer a glimpse onto common patterns of metabolic reprogramming across cancers, and the work serves as a large-scale resource accessible via a web application

Social stress can lead to the development of psychological problems ranging from exaggerated anxiety and depression to antisocial and violence-related behaviors. Increasing evidence suggests that the immune system is involved in responses to social stress in adulthood. For example, human studies show that individuals with high aggression traits display heightened inflammatory cytokine levels and dysregulated immune responses such as slower wound healing. Similar findings have been observed in patients with depression, and comorbidity of depression and aggression was correlated with stronger immune dysregulation. Therefore, dysregulation of the immune system may be one of the mediators of social stress that produces aggression and/or depression. Similar to humans, aggressive animals also show increased levels of several proinflammatory cytokines, however, unlike humans these animals are more protected frominfectious organisms and have faster wound healing than animals with low aggression. On the other hand, subordinate animals that receive repeated social defeat stress have been shown to develop escalated and dysregulated immune responses such as glucocorticoid insensitivity in monocytes. In this review we synthesize the current evidence in humans, non-human primates, and rodents to show a role for the immune system in responses to social stress leading to psychiatric problems such as aggression or depression. We argue that while depression and aggression represent two fundamentally different behavioral and physiological responses to social stress, it is possible that some overlapped, as well as distinct, pattern of immune signaling may underlie both of them. We also argue the necessity of studying animal models of maladaptive aggression induced by social stress (i.e., social isolation) for understanding neuro-immune mechanism of aggression, which may be relevant to human aggression.

Somatosensory neurons mediate responses to diverse mechanical stimuli, from innocuous touch to noxious pain. While recent studies have identified distinct populations of A mechanonociceptors (AMs) that are required for mechanical pain, the molecular underpinnings of mechanonociception remain unknown. Here, we show that the bioactive lipid sphingosine 1-phosphate (S1P) and S1P Receptor 3 (S1PR3) are critical regulators of acute mechanonociception. Genetic or pharmacological ablation of S1PR3, or blockade of S1P production, significantly impaired the behavioral response to noxious mechanical stimuli, with no effect on responses to innocuous touch or thermal stimuli. These effects are mediated by fast-conducting A mechanonociceptors, which displayed a significant decrease in mechanosensitivity in S1PR3 mutant mice. We show that S1PR3 signaling tunes mechanonociceptor excitability via modulation of KCNQ2/3 channels. Our findings define a new role for S1PR3 in regulating neuronal excitability and establish the importance of S1P/S1PR3 signaling in the setting of mechanical pain thresholds.

Laboratory animal programs and core laboratories often set service rates based on cost estimates. However, actual costs may be unknown, and service rates may not reflect the actual cost of services. Accurately evaluating the actual costs of services can be challenging and time-consuming. We used a time-driven activity-based costing (ABC) model to determine the cost of services provided by a resource laboratory at our institution. The time-driven approach is a more efficient approach to calculating costs than using a traditional ABC model. We calculated only 2 parameters: the time required to perform an activity and the unit cost of the activity based on employee cost. This method allowed us to rapidly and accurately calculate the actual cost of services provided, including microinjection of a DNA construct, microinjection of embryonic stem cells, embryo transfer, and in vitro fertilization. We successfully implemented a time-driven ABC model to evaluate the cost of these services and the capacity of labor used to deliver them. We determined how actual costs compared with current service rates. In addition, we determined that the labor supplied to conduct all services (10,645 min/wk) exceeded the practical labor capacity (8400 min/wk), indicating that the laboratory team was highly efficient and that additional labor capacity was needed to prevent overloading of the current team. Importantly, this time-driven ABC approach allowed us to establish a baseline model that can easily be updated to reflect operational changes or changes in labor costs. We demonstrated that a time-driven ABC model is a powerful management tool that can be applied to other core facilities as well as to entire animal programs, providing valuable information that can be used to set rates based on the actual cost of services and to improve operating efficiency.