The rockefeller university

We have reported previously that autophagy is responsible for amyloid precursor protein-C-terminal fragment (APP-CTF) degradation and therefore A clearance. To elucidate the underlying mechanism, using LC3 affinity purification and mass spectrometry analysis, immunoprecipitation (IP), as well as live imaging analysis, we identified and demonstrated that the adaptor-related protein complex 2 (AP2) and PICALM (phosphatidylinositol binding clathrin assembly protein) are in a complex with LC3 and APP-CTF. Taken together, this new set of data suggests that the AP2-PICALM complex functions as an autophagic cargo receptor for the recognition and shipment of APP-CTF from the endocytic pathway to the LC3-dependent autophagic degradation pathway. Interestingly this AP2-LC3 connection seems to be involved in chemically-induced APP-CTF clearance as we observed using the small compound SMER28. The effect observed following SMER28 was significantly reduced after silencing AP2. While more work is required to elucidate the detailed molecular mechanisms involved, our actual data suggest that there is some level of specificity in the steps mentioned above.

The Clinical and Translational Science Awards (CTSA) program represents a significant public investment. To realize its major goal of improving the public's health and reducing health disparities, the CTSA Consortium's Community Engagement Key Function Committee has undertaken the challenge of developing a taxonomy of community health indicators. The objective is to initiate a unified approach for monitoring progress in improving population health outcomes. Such outcomes include, importantly, the interests and priorities of community stakeholders, plus the multiple, overlapping interests of universities and of the public health and health care professions involved in the development and use of local health care indicators. The emerging taxonomy of community health indicators that the authors propose supports alignment of CTSA activities and facilitates comparative effectiveness research across CTSAs, thereby improving the health of communities and reducing health disparities. The proposed taxonomy starts at the broadest level, determinants of health; subsequently moves to more finite categories of community health indicators; and, finally, addresses specific quantifiable measures. To illustrate the taxonomy's application, the authors have synthesized 21 health indicator projects from the literature and categorized them into international, national, or local/special jurisdictions. They furthered categorized the projects within the taxonomy by ranking indicators with the greatest representation among projects and by ranking the frequency of specific measures. They intend for the taxonomy to provide common metrics for measuring changes to population health and, thus, extend the utility of the CTSA Community Engagement Logic Model. The input of community partners will ultimately improve population health.

HLA-restricted immune escape mutations that persist following HIV transmission could gradually spread through the viral population, thereby compromising host antiviral immunity as the epidemic progresses. To assess the extent and phenotypic impact of this phenomenon in an immunogenetically diverse population, we genotypically and functionally compared linked HLA and HIV (Gag/Nef) sequences from 358 historic (1979-1989) and 382 modern (2000-2011) specimens from four key cities in the North American epidemic (New York, Boston, San Francisco, Vancouver). Inferred HIV phylogenies were star-like, with approximately two-fold greater mean pairwise distances in modern versus historic sequences. The reconstructed epidemic ancestral (founder) HIV sequence was essentially identical to the North American subtype B consensus. Consistent with gradual diversification of a "consensus-like" founder virus, the median "background" frequencies of individual HLA-associated polymorphisms in HIV (in individuals lacking the restricting HLA[s]) were similar to 2-fold higher in modern versus historic HIV sequences, though these remained notably low overall (e. g. in Gag, medians were 3.7% in the 2000s versus 2.0% in the 1980s). HIV polymorphisms exhibiting the greatest relative spread were those restricted by protective HLAs. Despite these increases, when HIV sequences were analyzed as a whole, their total average burden of polymorphisms that were "pre-adapted" to the average host HLA profile was only similar to 2% greater in modern versus historic eras. Furthermore, HLA-associated polymorphisms identified in historic HIV sequences were consistent with those detectable today, with none identified that could explain the few HIV codons where the inferred epidemic ancestor differed from the modern consensus. Results are therefore consistent with slow HIV adaptation to HLA, but at a rate unlikely to yield imminent negative implications for cellular immunity, at least in North America. Intriguingly, temporal changes in protein activity of patient-derived Nef (though not Gag) sequences were observed, suggesting functional implications of population-level HIV evolution on certain viral proteins.

The production of (1S), (2S), and (3S) is investigated in pPb and pp collisions at centre-of-mass energies per nucleon pair of 5.02 TeV and 2.76 TeV, respectively. The datasets correspond to integrated luminosities of about 31 nb(-1) (pPb) and 5.4 pb(-1) (pp), collected in 2013 by the CMS experiment at the LHC. Upsilons that decay into muons are reconstructed within the rapidity interval |y (CM)| < 1.93 in the nucleon-nucleon centre-of-mass frame. Their production is studied as a function of two measures of event activity, namely the charged-particle multiplicity measured in the pseudorapidity interval |eta| < 2.4, and the sum of transverse energy deposited at forward pseudorapidity, 4.0 < |eta| < 5.2. The cross sections normalized by their event activity integrated values, (nS)/aEuro(nS)aEuro parts per thousand, are found to rise with both measures of the event activity in pp and pPb. In both collision systems, the ratios of the excited to the ground state cross sections, (nS)/ (1S), are found to decrease with the charged-particle multiplicity, while as a function of the transverse energy the variation is less pronounced. The event activity integrated double ratios, [(nS)/ (1S)](pPb) /[(nS)/ (1S)](pp), are also measured and found to be 0.83 +/- 0.05 (stat.) +/- 0.05 (syst.) and 0.71 +/- 0.08 (stat.) +/- 0.09 (syst.) for (2S) and (3S), respectively.

Background: IRAK4 is a central kinase in IL-1R/TLR signaling. Results: IRAK4 is activated by autophosphorylation, and its inhibition reduces cytokine induction in human monocytes but not dermal fibroblasts. Conclusion: IL-1R/TLR-induced autophosphorylation activates IRAK4 and controls cytokine induction in a cell type-specific manner. Significance: Our data provide the mechanism of IRAK4 activation and role in cytokine induction in human cells. IRAK4 is a central kinase in innate immunity, but the role of its kinase activity is controversial. The mechanism of activation for IRAK4 is currently unknown, and little is known about the role of IRAK4 kinase in cytokine production, particularly in different human cell types. We show IRAK4 autophosphorylation occurs by an intermolecular reaction and that autophosphorylation is required for full catalytic activity of the kinase. Phosphorylation of any two of the residues Thr-342, Thr-345, and Ser-346 is required for full activity, and the death domain regulates the activation of IRAK4. Using antibodies against activated IRAK4, we demonstrate that IRAK4 becomes phosphorylated in human cells following stimulation by IL-1R and Toll-like receptor agonists, which can be blocked pharmacologically by a dual inhibitor of IRAK4 and IRAK1. Interestingly, in dermal fibroblasts, although complete inhibition of IRAK4 kinase activity does not inhibit IL-1-induced IL-6 production, NF-B, or MAPK activation, there is complete ablation of these processes in IRAK4-deficient cells. In contrast, the inhibition of IRAK kinase activity in primary human monocytes reduces R848-induced IL-6 production with minimal effect on NF-B or MAPK activation. Taken together, these studies define the mechanism of IRAK4 activation and highlight the differential role of IRAK4 kinase activity in different human cell types as well as the distinct roles IRAK4 scaffolding and kinase functions play.

A search for the production of heavy partners of the top quark with charge 5/3 is performed in events with a pair of same-sign leptons. The data sample corresponds to an integrated luminosity of 19.5 fb(-1) and was collected at root s = 8 TeV by the CMS experiment. No significant excess is observed in the data above the expected background, and the existence of top-quark partners with masses below 800 GeV is excluded at a 95% confidence level, assuming they decay exclusively to tW. This is the first limit on these particles from the LHC, and it is significantly more restrictive than previous limits.

Although non-medical use of oxycodone continues to be a growing problem in the United States, there are no animal studies examining the effects of long-term oxycodone self-administration (SA). The current study was designed to examine chronic oxycodone SA by mice (14 days), in novel extended (4 h) SA sessions and its effect on selective striatal neurotransmitter receptor mRNA expression. Adult male C57/BL6J mice were either allowed to self-administer oxycodone (0.25 mg/kg/infusion, FR1) or served as yoked-saline controls in an extended access paradigm. Mice self-administered oxycodone for 4 h/day for 14 consecutive days. Comparison groups with 14-days exposure to 1-h SA sessions were also studied. Within 1 h of the last extended SA session, mice were sacrificed, dorsal striatum was isolated and selective neurotransmitter receptor mRNA levels were examined. The oxycodone groups poked the active hole significantly more times than the yoked controls. The number of nose pokes at the active hole rose over the 14 days in the oxycodone group with extended access. The expression of 13 neurotransmitter receptor mRNAs was significantly altered in the dorsal striatum, including the gamma-aminobutyric acid (GABA) A receptor beta 2 subunit (Gabrb2) showing experiment-wise significant decrease, as a result of extended oxycodone SA. C57BL/6 J mice escalated the amount of oxycodone self-administered across 14 consecutive daily extended sessions, but not 1-h sessions. Decreases in Gabrb2 mRNA levels may underlie escalation of oxycodone intake in the extended access SA sessions.

Coagulase-negative staphylococci (CoNS) are believed to function as reservoirs, as well as possible sources of staphylococcal chromosome cassette mec (SCCmec) to Staphylococcus aureus, but the frequency, preferred partners, and factors promoting SCCmec transfer are not known. Such postulated in vivo genetic transfer events are likely to occur at anatomical sites such as the normal nasal mucosa, which is known to be colonized by both CoNS and coagulase positive staphylococci. In this study, we characterized S. aureus and CoNS strains colonizing the anterior nares of 67 patients in Denmark. A total of 54 patients (80%) were colonized with staphylococci that included nine different species identified by internal transcribed spacer PCR (ITS-PCR) and 16S RNA sequencing. The highest rates of colonization were found for S. epidermidis (58%) and S. aureus (39%). Methicillin resistance was present in S. aureus (53%), S. epidermidis (53%), S. haemolyticus (33%), and S. hominis (62%). Genetic backgrounds were characterized by spa typing for S. aureus and by pulsed-field gel electrophoresis for CoNS. SCCmec typing showed that SCCmec type IV (2B) was the most common in the entire collection (65%). Carriage of multiple species was detected in 20 patients (30%), 16 of whom were colonized with both S. aureus and S. epidermidis. In two cases, simultaneous carriage of different methicillin resistant species was detected. However, the strains carried different SCCmec types. Additional studies in the same epidemiological settings are warranted to identify interspecific genetic events that involve the acquisition of SCCmec by S. aureus.

Langerhans cell histiocytosis (LCH) is a clonal disorder with elusive etiology, characterized by the accumulation of CD207(+) dendritic cells (DCs) in inflammatory lesions. Recurrent BRAF-V600E mutations have been reported in LCH. In this study, lesions from 100 patients were genotyped, and 64% carried the BRAF-V600E mutation within infiltrating CD207(+) DCs. BRAF-V600E expression in tissue DCs did not define specific clinical risk groups but was associated with increased risk of recurrence. Strikingly, we found that patients with active, high-risk LCH also carried BRAF-V600E in circulating CD11c(+) and CD14(+) fractions and in bone marrow (BM) CD34(+) hematopoietic cell progenitors, whereas the mutation was restricted to lesional CD207(+) DC in low-risk LCH patients. Importantly, BRAF-V600E expression in DCs was sufficient to drive LCH-like disease in mice. Consistent with our findings in humans, expression of BRAF-V600E in BM DC progenitors recapitulated many features of the human high-risk LCH, whereas BRAF-V600E expression in differentiated DCs more closely resembled low-risk LCH. We therefore propose classification of LCH as a myeloid neoplasia and hypothesize that high-risk LCH arises from somatic mutation of a hematopoietic progenitor, whereas low-risk disease arises from somatic mutation of tissue-restricted precursor DCs.

Faithful chromosome segregation during meiosis is indispensable to prevent birth defects and infertility. Canonical genetic manipulations have not been very useful for studying meiosis II, since mutations of genes involved in cell cycle regulation or chromosome segregation may affect meiosis I, making interpretations of any defects observed in meiosis II complicated. Here we present a powerful strategy to dissect meiosis I and meiosis II, using chemical inhibitors in genetically tractable model organism fission yeast (Schizosaccharomyces pombe). As various chemical probes are not active in fission yeast, mainly due to an effective multidrug resistance (MDR) response, we have recently developed a drug-hypersensitive MDR-sup strain by suppression of the key genes responsible for MDR response. We further developed the MDR-supML (marker-less) strain by deleting 7 MDR genes without commonly used antibiotic markers. The new strain makes fluorescent tagging and gene deletion much simpler, which enables effective protein visualization in varied genetic backgrounds. Using the MDR-supML strain with chemical inhibitors and live cell fluorescence microscopy, we established cell cycle arrest at meiosis I and meiosis II and examined Aurora-dependent spindle assembly checkpoint (SAC) regulation during meiosis. We found that Aurora B/Ark1 kinase activity is required for recruitment of Bub1, an essential SAC kinase, to unattached kinetochore in prometaphase I and prometaphase II as in mitosis. Thus, Aurora's role in SAC activation is likely conserved in mitosis, meiosis I, and meiosis II. Together, our MDR-supML strain will be useful to dissect complex molecular mechanisms in mitosis and 2 successive meiotic divisions.