The rockefeller university

Following their synthesis in the endoplasmic reticulum (ER), voltage-gated sodium channels (Na-V) are transported to the membranes of excitable cells, where they often cluster, such as at the axon initial segment of neurons. Although the mechanisms by which Na-V channels form and maintain clusters have been extensively examined, the processes that govern their transport and degradation have received less attention. Our entry into the study of these processes began with the isolation of a new allele of the zebrafish mutant alligator, which we found to be caused by mutations in the gene encoding really interesting new gene (RING) finger protein 121 (RNF121), an E3-ubiquitin ligase present in the ER and cis-Golgi compartments. Here we demonstrate that RNF121 facilitates two opposing fates of Na-V channels: (i) ubiquitin-mediated proteasome degradation and (ii) membrane localization when coexpressed with auxiliary Na-V beta subunits. Collectively, these results indicate that RNF121 participates in the quality control of NaV channels during their synthesis and subsequent transport to the membrane.

The production of Z bosons is studied in the dimuon and dielectron decay channels in PbPb and pp collisions at root s(NN) = 2.76TeV, using data collected by the CMS experiment at the LHC. The PbPb data sample corresponds to an integrated luminosity of about 166 mu b(-1), while the pp data sample collected in 2013 at the same nucleon-nucleon centre-of-mass energy has an integrated luminosity of 5.4 pb(-1). The Z boson yield is measured as a function of rapidity, transverse momentum, and collision centrality. The ratio of PbPb to pp yields, scaled by the number of inelastic nucleon-nucleon collisions, is found to be 1.06 +/- 0.05 (stat) +/- 0.08 (syst) in the dimuon channel and 1.02 +/- 0.08 (stat) +/- 0.15 (syst) in the dielectron channel, for centrality-integrated Z boson production. This binary collision scaling is seen to hold in the entire kinematic region studied, as expected for a colourless probe that is unaffected by the hot and dense QCD medium produced in heavy ion collisions.

Dissecting and studying cellular systems requires the ability to specifically isolate distinct proteins along with the co-assembled constituents of their associated complexes. Affinity capture techniques leverage high affinity, high specificity reagents to target and capture proteins of interest along with specifically associated proteins from cell extracts. Affinity capture coupled to mass spectrometry (MS)-based proteomic analyses has enabled the isolation and characterization of a wide range of endogenous protein complexes. Here, we outline effective procedures for the affinity capture of protein complexes, highlighting best practices and common pitfalls.

Long-term in vivo expression of a broad and potent entry inhibitor could circumvent the need for a conventional vaccine for HIV-1. Adeno-associated virus (AAV) vectors can stably express HIV-1 broadly neutralizing antibodies (bNAbs)(1,2). However, even the best bNAbs neutralize 10-50% of HIV-1 isolates inefficiently (80% inhibitory concentration (IC80) > 5 mu g ml(-1)), suggesting that high concentrations of these antibodies would be necessary to achieve general protection(3-6). Here we show that eCD4-Ig, a fusion ofCD4-Ig with a small CCR5-mimetic sulfopeptide, binds avidly and cooperatively to the HIV-1 envelope glycoprotein (Env) and is more potent than the best bNAbs (geometric mean half-maximum inhibitory concentration (IC50), 0.05 mu g ml(-1)). Because eCD4-Ig binds only conserved regions of Env, it is also much broader than any bNAb. For example, eCD4-Ig efficiently neutralized 100% of a diverse panel of neutralization-resistant HIV-1, HIV-2 and simian immunodeficiency virus isolates, including a comprehensive set of isolates resistant to the CD4-binding site bNAbs VRC01, NIH45-46 and 3BNC117. Rhesus macaques inoculated with an AAV vector stably expressed 17-77 mu g ml(-1) of fully functional rhesuseCD4-Ig for more than 40 weeks, and these macaques were protected from several infectious challenges with SHIV-AD8. Rhesus eCD4-Ig was also markedly less immunogenic than rhesus forms of four well-characterized bNAbs. Our data suggest that AAV-delivered eCD4-Ig can function like an effective HIV-1 vaccine.

Two subsets of conventional dendritic cells (cDCs) with distinct cell surface markers and functions exist in mouse and human. The two subsets of cDCs are specialized antigen-presenting cells that initiate T cell immunity and tolerance. In the mouse, a migratory cDC precursor (pre-CDC) originates from defined progenitors in the bone marrow (BM). Small numbers of short-lived pre-CDCs travel through the blood and replace cDCs in the peripheral organs, maintaining homeostasis of the highly dynamic cDC pool. However, the identity and distribution of the immediate precursor to human cDCs has not been defined. Using a tissue culture system that supports the development of human DCs, we identify a migratory precursor (hpre-CDC) that exists in human cord blood, BM, blood, and peripheral lymphoid organs. hpre-CDCs differ from premonocytes that are restricted to the BM. In contrast to earlier progenitors with greater developmental potential, the hpre-CDC is restricted to producing CD1c(+) and CD141(+) Clec9a(+) cDCs. Studies in human volunteers demonstrate that hpre-CDCs are a dynamic population that increases in response to levels of circulating Flt3L.

Studying 830 pre-B ALL cases from four clinical trials, we found that human ALL can be divided into two fundamentally distinct subtypes based on pre-BCR function. While absent in the majority of ALL cases, tonic pre-BCR signaling was found in 112 cases (13.5%). In these cases, tonic pre-BCR signaling induced activation of BCL6, which in turn increased pre-BCR signaling output at the transcriptional level. Interestingly, inhibition of pre-BCR-related tyrosine kinases reduced constitutive BCL6 expression and selectively killed patient-derived pre-BCR+ ALL cells. These findings identify a genetically and phenotypically distinct subset of human ALL that critically depends on tonic pre-BCR signaling. In vivo treatment studies suggested that pre-BCR tyrosine kinase inhibitors are useful for the treatment of patients with pre-BCR+ ALL.

Recent X-ray crystallographic studies of Pol II in complex with the general transcription factor (GTF) IIB have begun to provide insights into the mechanism of transcription initiation. These structures have also shed light on the architecture of the transcription preinitiation complex (PIC). However, structural characterization of a functional PIC is still lacking, and even the topological arrangement of the GTFs in the Pol II complex is a matter of contention. We have extended our activity-based affinity crosslinking studies, initially developed to investigate the interaction of bacterial RNA polymerase with s, to the eukaryotic transcription machinery. Towards that end, we sought to identify GTFs that are within the Pol II active site in a functioning PIC. We provide biochemical evidence that TFIIB is located within similar to 9 angstrom of the -2 site of promoter DNA, where it is positioned to play a role in de novo transcription initiation.

Electronics companies throughout Asia recognize the benefits of Green Supply Chain Management (GSCM) for gaining competitive advantage. A large majority of electronics companies in Taiwan have recently adopted the Electronic Industry Citizenship Coalition (EICC) Code of Conduct for defining and managing their social and environmental responsibilities throughout their supply chains. We surveyed 106 Tier 1 suppliers to the Taiwanese computer industry to determine their environmental performance using the EICC Code of Conduct (EICC Code) and performed Analysis of Variance (ANOVA) on the 63/106 questionnaire responses collected. We test the results to determine whether differences in product type, geographic area, and supplier size correlate with different levels of environmental performance. To our knowledge, this is the first study to analyze questionnaire data on supplier adoption to optimize the implementation of GSCM. The results suggest that characteristic classification of suppliers could be employed to enhance the efficiency of GSCM.

Recent advances in sequencing technologies have greatly enhanced our abilities to identify novel microbial sequences. Thus, our understanding of the global virome and the virome of specific host species in particular is rapidly expanding. Identification of animal viruses is important for understanding animal disease, the origin and evolution of human viruses, as well as zoonotic reservoirs for emerging infections. Although the human hepacivirus, hepatitis C virus (HCV), was identified 25 years ago, its origin has remained elusive. In 2011, the first HCV homolog was reported in dogs but subsequent studies showed the virus to be widely distributed in horses. This indicated a wider hepacivirus host range and paved the way for identification of rodent, bat and non-human primate hepaciviruses. The equine non-primate hepacivirus (NPHV) remains the closest relative of HCV and is so far the best characterized. Identification and characterization of novel hepaciviruses may in addition lead to development of tractable animal models to study HCV persistence, immune responses and pathogenesis. This could be particular important, given the current shortage of immunocompetent models for robust HCV infection. Much remains to be learned on the novel hepaciviruses, including their association with disease, and thereby how relevant they will become as HCV model systems and for studies of animal disease. This review discusses how virome analysis led to identification of novel hepaci- and pegiviruses, their genetic relationship and characterization and the potential use of animal hepaciviruses as models to study hepaciviral infection, immunity and pathogenesis. This article forms part of a symposium in Antiviral Research on "Hepatitis C: Next steps toward global eradication." (C) 2015 Elsevier B.V. All rights reserved.

Primordial germ cells (PGCs) are the earliest population of germ cells established during embryonic development and constitute the beginning of the totipotent state. A recent study provides a new protocol for the efficient generation of PGC-like cells from human embryonic stem cells, providing an in vitro platform to study human PGC differentiation and specification.