T cell signatures and modulation by regulatory T cells

CD4 T cells are crucial coordinators of protective immune responses against microbes and tumors. However, immune responses misdirected towards self-antigens are the hallmarks of autoimmune diseases. Regulatory T cells (Tregs) maintain peripheral immune tolerance, mainly by suppressing effector T cells. Unfortunately, immunosuppression by Tregs can also favor cancer. Regulating the susceptibility of T cells towards Tregmeditated immunosuppression presents an exciting approach to reconcile between immune activation and tolerance in a disease-specific way. However, less is known about the regulation and mechanism of Treg-mediated suppression in target T cells. The work presented in this thesis is dedicated to unraveling the modulation of signaling cascades in T cells upon T cell receptor (TCR) stimulation and suppression by Tregs. By using unbiased phosphoproteomics, we have mapped the global phosphoproteome of T cells upon TCR stimulation and suppression by Tregs. Our data indicate that Tregs suppress T cells mainly by opposing activation-induced phosphorylation. We discovered that Tregs revert activation-induced phosphorylation of DEF6 at specific sites (T 595 / S 597). Utilizing phospho-mutants, we discovered novel functions of these phosphorylations in disrupting the interaction of DEF6 with the IP3R and regulating T cell signaling via modulation of NFAT activation and transcriptional regulation of T cell cytokines. Upon further exploration of the phosphoproteomic candidate list, we observed similarly altered phospho-regulation of protein phosphatase 1, regulatory subunit 11 (PPP1R11) by Tregs. We revealed that silencing of PPP1R11 renders T cells resistant to Treg-mediation suppression. Our data indicates PPP1R11 to be a novel regulator of T cell activation and Treg-mediated suppression. In light of several reports on resistant T cells in multiple diseases, the phosphoproteomic mapping of suppressed T cells and the insights into novel roles of DEF6 and PPP1R11 from our study may aid in regulating the sensitivity of T cells towards immunosuppression by Tregs in cancer and autoimmune diseases. Besides phosphorylation of proteins, the subcellular localization of proteins is also an important regulatory mechanism of protein activity. Protein localization and activity is modulated by TCR stimulation and was shown for well-known TCR signaling proteins such as NFAT to be affected by Tregs as well. However, these mechanisms are not understood on a global level. Since there is no global data set available on subcellular protein localization and TCR stimulation-induced translocation in primary human T cells, as the first milestone in this field, we have performed a comprehensive mapping of the spatial proteome of T cells and TCR-induced subcellular protein translocation. Besides exploring the target T cell side, in this work we also studied novel methods to induce Tregs, which represents the “other side” of Treg-based immune therapies. We present a novel method of inducing iTregs by using supernatants from M2 macrophages. Our data demonstrate that M2 macrophages induce iTregs by binding and re-releasing TGF-β, which may be explored for Treg induction in situ in the future. Together, by presenting a global picture of T cell protein signaling yet with fine resolution, our work provides new mechanisms and data sets to revisit the role of T cells in therapy, especially in the context of T cell suppression by Tregs

Analysis of KatG Ser315Thr Mutation in Multidrug Resistant Mycobacterium tuberculosis and SLC11A1 Polymorphism in Multidrug Resistance Tuberculosis in Central Development Region of Nepal Using PCR-RFLP Technique: A Pilot Study

Ser315Thr mutations in genes encoding the mycobacteria catalase-peroxidase (KatG) has been associated with the major resistance to isoniazid (INH) in Mycobacterium tuberculosis (MTB). Also G/C polymorphisms in INT4 region of the solute carrier family 11 member 1 gene (SLC11A1) and susceptibility towards tuberculosis (TB) has been demonstrated worldwide. 24 drug resistant MTB culture positive samples and 24 whole?blood samples were collected from different TB patients of Central Development Region of Nepal in 2009. A Polymerase Chain Reaction (PCR) - Restriction Fragment Length Polymorphism (RFLP) assay was carried out in order to investigate Ser315Thr KatG mutation and G/C polymorphism in INT4 region. 4 (16.67%) samples out of 24 MTB culture samples demonstrated the Ser315Thr KatG mutation whereas none of the 24 whole blood samples were found to contain G/C polymorphism in INT4. Though no significant correlation could be found between INT4 polymorphism and TB susceptibility, overall scenario of Nepal cannot be drawn from this data. Molecular diagnostic technique such as PCR-RFLP can be used in a robust scale to carry out base line studies in the TB population of Nepal. Key words: Multi?drug resistance; Tuberculosis; PCR; RFLP Nepal Journal of Biotechnology. Jan. 2011, Vol. 1, No. 1 : 14-2

Non-parametric combination analysis of multiple data types enables detection of novel regulatory mechanisms in T cells of multiple sclerosis patients

Multiple Sclerosis (MS) is an autoimmune disease of the central nervous system with prominent neurodegenerative components. The triggering and progression of MS is associated with transcriptional and epigenetic alterations in several tissues, including peripheral blood. The combined influence of transcriptional and epigenetic changes associated with MS has not been assessed in the same individuals. Here we generated paired transcriptomic (RNA-seq) and DNA methylation (Illumina 450 K array) profiles of CD4+ and CD8+ T cells (CD4, CD8), using clinically accessible blood from healthy donors and MS patients in the initial relapsing-remitting and subsequent secondary-progressive stage. By integrating the output of a differential expression test with a permutation-based non-parametric combination methodology, we identified 149 differentially expressed (DE) genes in both CD4 and CD8 cells collected from MS patients. Moreover, by leveraging the methylation-dependent regulation of gene expression, we identified the gene SH3YL1, which displayed significant correlated expression and methylation changes in MS patients. Importantly, silencing of SH3YL1 in primary human CD4 cells demonstrated its influence on T cell activation. Collectively, our strategy based on paired sampling of several cell-types provides a novel approach to increase sensitivity for identifying shared mechanisms altered in CD4 and CD8 cells of relevance in MS in small sized clinical materials

Measurements of the W production cross sections in association with jets with the ATLAS detector

This paper presents cross sections for the production of a W boson in association with jets, measured in proton–proton collisions at √s=7 TeV with the ATLAS experiment at the large hadron collider. With an integrated luminosity of 4.6 fb−1, this data set allows for an exploration of a large kinematic range, including jet production up to a transverse momentum of 1 TeV and multiplicities up to seven associated jets. The production cross sections for W bosons are measured in both the electron and muon decay channels. Differential cross sections for many observables are also presented including measurements of the jet observables such as the rapidities and the transverse momenta as well as measurements of event observables such as the scalar sums of the transverse momenta of the jets. The measurements are compared to numerous QCD predictions including next-to-leading-order perturbative calculations, resummation calculations and Monte Carlo generators

Socializing One Health: an innovative strategy to investigate social and behavioral risks of emerging viral threats

In an effort to strengthen global capacity to prevent, detect, and control infectious diseases in animals and people, the United States Agency for International Development’s (USAID) Emerging Pandemic Threats (EPT) PREDICT project funded development of regional, national, and local One Health capacities for early disease detection, rapid response, disease control, and risk reduction. From the outset, the EPT approach was inclusive of social science research methods designed to understand the contexts and behaviors of communities living and working at human-animal-environment interfaces considered high-risk for virus emergence. Using qualitative and quantitative approaches, PREDICT behavioral research aimed to identify and assess a range of socio-cultural behaviors that could be influential in zoonotic disease emergence, amplification, and transmission. This broad approach to behavioral risk characterization enabled us to identify and characterize human activities that could be linked to the transmission dynamics of new and emerging viruses. This paper provides a discussion of implementation of a social science approach within a zoonotic surveillance framework. We conducted in-depth ethnographic interviews and focus groups to better understand the individual- and community-level knowledge, attitudes, and practices that potentially put participants at risk for zoonotic disease transmission from the animals they live and work with, across 6 interface domains. When we asked highly-exposed individuals (ie. bushmeat hunters, wildlife or guano farmers) about the risk they perceived in their occupational activities, most did not perceive it to be risky, whether because it was normalized by years (or generations) of doing such an activity, or due to lack of information about potential risks. Integrating the social sciences allows investigations of the specific human activities that are hypothesized to drive disease emergence, amplification, and transmission, in order to better substantiate behavioral disease drivers, along with the social dimensions of infection and transmission dynamics. Understanding these dynamics is critical to achieving health security--the protection from threats to health-- which requires investments in both collective and individual health security. Involving behavioral sciences into zoonotic disease surveillance allowed us to push toward fuller community integration and engagement and toward dialogue and implementation of recommendations for disease prevention and improved health security

Search for new phenomena in events with three or more charged leptons in pppp collisions at s=8\sqrt{s}=8 TeV with the ATLAS detector

A generic search for anomalous production of events with at least three charged leptons is presented. The data sample consists of $pp$ collisions at $\sqrt{s}=8$ TeV collected in 2012 by the ATLAS experiment at the CERN Large Hadron Collider, and corresponds to an integrated luminosity of 20.3 fb$^{-1}$. Events are required to have at least three selected lepton candidates, at least two of which must be electrons or muons, while the third may be a hadronically decaying tau. Selected events are categorized based on their lepton flavour content and signal regions are constructed using several kinematic variables of interest. No significant deviations from Standard Model predictions are observed. Model-independent upper limits on contributions from beyond the Standard Model phenomena are provided for each signal region, along with prescription to re-interpret the limits for any model. Constraints are also placed on models predicting doubly charged Higgs bosons and excited leptons. For doubly charged Higgs bosons decaying to $e\tau$ or $\mu\tau$, lower limits on the mass are set at 400 GeV at 95% confidence level. For excited leptons, constraints are provided as functions of both the mass of the excited state and the compositeness scale $\Lambda$, with the strongest mass constraints arising in regions where the mass equals $\Lambda$. In such scenarios, lower mass limits are set at 3.0 TeV for excited electrons and muons, 2.5 TeV for excited taus, and 1.6 TeV for every excited-neutrino flavour.Comment: 442 pages plus author list + cover pages (60 pages total), 6 figures, 25 tables, submitted to JHEP, All figures including auxiliary figures are available at http://atlas.web.cern.ch/Atlas/GROUPS/PHYSICS/PAPERS/EXOT-2012-20

Non-parametric combination analysis of multiple data types enables detection of novel regulatory mechanisms in T cells of multiple sclerosis patients.

Multiple Sclerosis (MS) is an autoimmune disease of the central nervous system with prominent neurodegenerative components. The triggering and progression of MS is associated with transcriptional and epigenetic alterations in several tissues, including peripheral blood. The combined influence of transcriptional and epigenetic changes associated with MS has not been assessed in the same individuals. Here we generated paired transcriptomic (RNA-seq) and DNA methylation (Illumina 450 K array) profiles of CD4+ and CD8+ T cells (CD4, CD8), using clinically accessible blood from healthy donors and MS patients in the initial relapsing-remitting and subsequent secondary-progressive stage. By integrating the output of a differential expression test with a permutation-based non-parametric combination methodology, we identified 149 differentially expressed (DE) genes in both CD4 and CD8 cells collected from MS patients. Moreover, by leveraging the methylation-dependent regulation of gene expression, we identified the gene SH3YL1, which displayed significant correlated expression and methylation changes in MS patients. Importantly, silencing of SH3YL1 in primary human CD4 cells demonstrated its influence on T cell activation. Collectively, our strategy based on paired sampling of several cell-types provides a novel approach to increase sensitivity for identifying shared mechanisms altered in CD4 and CD8 cells of relevance in MS in small sized clinical materials