DOCSLIB.ORG

Inventories of Naive and Tolerant Mouse CD4 T Cell Repertoires Reveal a Hierarchy of Deleted and Diverted T Cell Receptors

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Inventories of Naive and Tolerant Mouse CD4 T Cell Repertoires Reveal a Hierarchy of Deleted and Diverted T Cell Receptors Inventories of naive and tolerant mouse CD4 T cell repertoires reveal a hierarchy of deleted and diverted T cell receptors Tobias Hasslera, Emanuel Urmanna, Sebastian Teschnera, Christine Federlea, Thamotharampillai Dileepanb, Kilian Schoberc, Marc K. Jenkinsb, Dirk H. Buschc,d, Maria Hinterbergera,1, and Ludger Kleina,1,2 aInstitute for Immunology, Faculty of Medicine, Ludwig-Maximilians-University, 82152 Planegg-Martinsried, Germany; bCenter for Immunology, University of Minnesota, Minneapolis, MN 55455; cInstitute for Medical Microbiology, Immunology and Hygiene, Technische Universität München, 81675 Munich, Germany; and dGerman Center for Infection Research, 81675 Munich, Germany Edited by Shimon Sakaguchi, WPI Immunology Frontier Research Center, Osaka University, Osaka, Japan, and approved August 5, 2019 (received for review May 2, 2019) – Deletion or Treg cell differentiation are alternative fates of autore- deletional tolerance (14 17), also shapes the Treg cell repertoire active MHCII-restricted thymocytes. How these different modes (18–20). of tolerance determine the size and composition of polyclonal co- MHCII tetramers were employed to trace minute cohorts of horts of autoreactive T cells with shared specificity is poorly un- CD4 T cells when their cognate antigens were transgenically derstood. We addressed how tolerance to a naturally expressed expressed to emulate a ubiquitous or TRA-like expression pat- autoantigen of the central nervous system shapes the CD4 T cell tern (21, 22). Widespread antigen expression was associated with repertoire. Specific cells in the tolerant peripheral repertoire either deletion, whereas a more restricted TRA-like expression ensued + + were Foxp3 or displayed anergy hallmarks and, surprisingly, in the emergence of Foxp3 Treg cells. While these findings were at least as frequent as in the nontolerant repertoire. Despite suggested that the mechanism of tolerance is somehow dictated this apparent lack of deletional tolerance, repertoire inventories by the antigen expression pattern, it remains to be seen whether uncovered that some T cell receptors (TCRs) were lost from the this is a generally applicable rule (23). CD4 T cell pool, whereas others mediated T cell differentiation. INFLAMMATION reg TCR transgenics, global repertoire sequencing, and MHC IMMUNOLOGY AND The antigen responsiveness of these TCRs supported an affinity tetramers each entail distinct limitations. Monoclonal systems model of central tolerance. Importantly, the contribution of differ- with unphysiologically high frequencies of antigen-specific cells ent diverter TCRs to the nascent thymic Treg cell population may not faithfully recapitulate T cell fate decisions in polyclonal reflected their antigen reactivity rather than their frequency settings (24, 25). Large-scale repertoire analyses are limited in among precursors. This reveals a multilayered TCR hierarchy in their potential to relate cell fate to antigen specificity. MHC CD4 T cell tolerance that separates deleted and diverted TCRs tetramers may fail to reveal holes in the tolerant repertoire, as and assures that the Treg cell compartment is filled with cells of the loss of certain TCRs might be numerically compensated by maximal permissive antigen reactivity. expansion of cells carrying others TCRs. Moreover, although the + + emergence of Foxp3 cells among Tetramer (Tet) cells indi- T cell tolerance | clonal deletion | clonal diversion | regulatory T cell | cates that T cell induction occurs, it remains unclear whether MHC class II tetramer reg this selectively applies to cells bearing distinct TCRs. cell receptor (TCR) stimulation of thymocytes can result in Significance Ttheir deletion but can also specify the diversion of MHCII- + restricted T cells into the Foxp3 regulatory T (T ) cell lineage reg Central tolerance can generate holes in the CD4 T cell repertoire (1). The determinants that specify these alternative cell fates are + or divert cells into the Foxp3 T regulatory (T )celllineage. incompletely understood, and little is known about how clonal reg Little is known with regard to how these diametrically different deletion and clonal diversion shape the composition of poly- cell fate decisions of autoreactive cells shape the size and com- clonal cohorts of autoreactive cells of shared antigen specificity. position of polyclonal cohorts of antigen-specific T cells. Here we TCR/model antigen transgenic systems showed that Treg cell generated inventories of very rare autoreactive T cells in the differentiation, similar to clonal deletion, can be instructed by naive versus tolerant polyclonal repertoire and identified T cell TCR agonists (2, 3), and this was later confirmed for polyclonal receptors (TCRs) that were lost, whereas others mediated Treg T cells and natural self-antigens (4). In some models, low antigen cell differentiation. The antigen responsiveness of these TCRs levels favored Treg cell generation, whereas high antigen doses revealed a TCR hierarchy that not only separates deleted from favored deletion, suggesting that Treg cell differentiation occurs diverted TCRs but also generates a Treg cell compartment with within an avidity window between positive selection and deletion high antigen reactivity. (5–8). Other observations, however, are difficult to reconcile with a reductionist, purely signal strength-based model. For in- Author contributions: M.H. and L.K. designed research; T.H., E.U., S.T., C.F., and M.H. performed research; T.D., K.S., M.K.J., and D.H.B. contributed new reagents/analytic tools; stance, concomitant deletion is seen in essentially all TCR T.H., E.U., C.F., M.H., and L.K. analyzed data; and L.K. wrote the paper. transgenic models of neoantigen-driven Treg cell diversion (2, 3, The authors declare no conflict of interest. 7, 9), and it remains possible that Treg cell differentiation entails This article is a PNAS Direct Submission. stochastic/selective elements (10). This open access article is distributed under Creative Commons Attribution-NonCommercial- Global TCR repertoire analyses supported the idea that Treg NoDerivatives License 4.0 (CC BY-NC-ND). differentiation is instructed by self-antigen recognition, although 1M.H. and L.K. contributed equally to this work. – some investigators came to opposing conclusions (11 13). Rep- 2To whom correspondence may be addressed. Email: [email protected]. ertoire analyses also indicated that Aire-dependent expression of This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. tissue-restricted antigens (TRAs) in thymic epithelial cells 1073/pnas.1907615116/-/DCSupplemental. (TECs), a phenomenon that had previously been associated with Published online August 26, 2019. www.pnas.org/cgi/doi/10.1073/pnas.1907615116 PNAS | September 10, 2019 | vol. 116 | no. 37 | 18537–18543 Downloaded by guest on September 26, 2021 Here we addressed how clonal deletion and Treg induction However, it remained possible that clonal deletion also shaped impact not only the size but also the TCR composition of poly- the composition of PLP-specific cells, yet was numerically clonal CD4 T cells recognizing the natural CNS autoantigen masked, for instance, by compensatory expansion of other PLP myelin proteolipid protein (PLP). BL/6 mice are largely resistant reactive cells. Distinguishing these possibilities requires a com- + to PLP-induced experimental autoimmune encephalomyelitis prehensive comparison of TCRs of Tet cells in Plp1KO versus (EAE), suggesting a robust state of antigen-specific tolerance. In Plp1WT mice. However, the sheer number of TCR α- and β-chain line with this, immunization of Plp1KO mice, to which PLP is a combinations that recognize a given antigen limits the feasibility foreign antigen, elicits a CD4 T cell response against 2 epitopes of such an approach in fully polyclonal repertoires. Several spanning amino acids 11 to 19 and 240 to 248, whereas Plp1WT hundred naive cells specific for a foreign antigen each expressed mice are functionally tolerant to these epitopes (26, 27). We a different TCR (29, 30). It is likely that the same applies to PLP- + assessed how far this unresponsiveness of the censored reper- specific CD4 T cells. Together with the paucity of Tet cells, this toire reflected the deletion, Treg cell diversion, or anergy of PLP- poses a significant hurdle to a conclusive comparison of antigen- specific CD4 T cells and whether these tolerance modes selec- specific TCRs in the absence or presence of a given autoantigen. tively applied to cells carrying distinct TCRs. To circumvent these inherent limitations of fully polyclonal TCR repertoires, we introduced a transgenic TCR β chain de- Results rived from a PLP11–19–specific TCR (27). The rationale was to PLP-specific CD4 T Cells Are Not Reduced in the Tolerant Repertoire reduce the repertoire complexity to the diversity of TCRα + but Contain Foxp3 Treg Cells. Using a combination of tetramer rearrangements and impose a bias toward higher numbers of staining and magnetic enrichment (SI Appendix, Fig. S1) (28), we PLP11–19–specific cells. The genotype of these compound trans- β TG Tcra+/− Foxp3GFP enumerated PLP11–19 (Tet-1) or PLP240–248 (Tet-3) specific CD4 genic mice (TCR -PLP1 :: :: ) will in the fol- T cells in pooled spleen and lymph node cells of Plp1KO and lowing be referred to as “Fixed-β.” Plp1WT mice. The uncensored peripheral repertoire of Plp1KO As expected, essentially all T cells in Fixed-β mice expressed + + mice contained 14.0 ± 3.0 Tet-1 CD4 T cells and
Load more

Recommended publications
  • Germ-Line Regulation of the Caenorhabditis Elegans Sex-Determining Gene Tra-2
    DEVELOPMENTAL BIOLOGY 204, 251–262 (1998) ARTICLE NO. DB989062 Germ-Line Regulation of the Caenorhabditis elegans Sex-Determining Gene tra-2 Patricia E. Kuwabara,* Peter G. Okkema,† and Judith Kimble‡ *MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, United Kingdom; †Laboratory for Molecular Biology, University of Illinois at Chicago, Chicago, Illinois 60607; and ‡Howard Hughes Medical Institute, Laboratory of Molecular Biology, Department of Biochemistry, and Department of Medical Genetics, University of Wisconsin, Madison, Wisconsin 53706 The Caenorhabditis elegans sex-determining gene tra-2 promotes female development of the XX hermaphrodite soma and germ line. We previously showed that a 4.7-kb tra-2 mRNA, which encodes the membrane protein TRA-2A, provides the primary feminizing activity of the tra-2 locus. This paper focuses on the germ-line activity and regulation of tra-2. First, we characterize a 1.8-kb tra-2 mRNA, which is hermaphrodite-specific and germ-line-dependent. This mRNA encodes TRA-2B, a protein identical to a predicted intracellular domain of TRA-2A. We show that the 1.8-kb mRNA is oocyte-specific, suggesting that it is involved in germ-line or embryonic sex determination. Second, we identify a tra-2 maternal effect on brood size that may be associated with the 1.8-kb mRNA. Third, we investigate seven dominant tra-2(mx) (for mixed character) mutations that sexually transform hermaphrodites to females by eliminating hermaphrodite spermatogenesis. Each of the tra-2(mx) mutants possesses a nonconserved missense change in a 22-amino-acid region common to both TRA-2A and TRA-2B, called the MX region.
    [Show full text]
  • And Crry, the Two Genetic Homologues of Human CR1 by Hector Molina,* Winnie Wong,~ Taroh Kinoshita,$ Carol Brenner,* Sharon Foley,* and V
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by PubMed Central Disfin_ct Receptor and Regulatory Properties of Recombinant Mouse Complement Receptor 1 (CR1) and Crry, the Two Genetic Homologues of Human CR1 By Hector Molina,* Winnie Wong,~ Taroh Kinoshita,$ Carol Brenner,* Sharon Foley,* and V. Michael Holers* From the *Howard Hughes Medical Institute Laboratories and Department of Medicine, Division of Rheumatology, Washington University School of Medicine, St. Louis, Missouri 63110; the *BASF Bioresearch Corporation, Cambridge, Massachusetts 02139; and the SDepartraent of Immunoregulation, Research Institute for Microbial Diseases, Osaka University, Osaka 565, Japan Summary The relationship between the characterized mouse regulators of complement activation (RCA) genes and the 190-kD mouse complement receptor 1 (MCK1), 155-kD mouse complement receptor 2 (MCR2), and mouse p65 is unclear. One mouse RCA gene, designatedMCR2 (or Cr2), encodes alternatively spliced 21 and 15 short consensus repeat (SCR)-containing transcripts that crosshybridize with cDNAs of both human CR2 and CR1, or CR2 alone, respectively. A five SCR-containing transcript derived from a second unique gene, designated Crry, also crosshybridizes with human CR1. We have previously shown that the 155-kD MCR2 is encoded by the 15 SCR-containing transcript. To analyze the protein products of the other transcripts, which are considered the genetic homologues of human CRI, we have expressed the 21 and the 5 SCR-containing cDNAs in the human K562 erythroleukemia cell line. We demonstrate that cells expressing the 21 SCK transcript express the 190-kD MCR1 protein. These cells react with five unique rat anti-MCR1 monodonal antibodies, including the 8C12 antibody considered to be monospecific for MCK1.
    [Show full text]
  • Cd79a Percp-Cy5.5 Noto Anche Come: Mb-1 Catalog Number(S): 9045-0792-025 (25 Tests), 9045-0792-120 (120 Tests)
    Page 1 of 2 CD79a PerCP-Cy5.5 Noto anche come: mb-1 Catalog Number(s): 9045-0792-025 (25 tests), 9045-0792-120 (120 tests) Profili di fluorescenza di normali linfociti del sangue periferico umano non colorati (istogramma blu) o colorati a livello intracellulare con CD79a coniugati con PerCP-Cy5.5 (istogramma viola). Informazioni sul prodotto Indice: CD79a PerCP-Cy5.5 Storage Conditions: Conservare a 2-8 °C. Catalog Number(s): 9045-0792-025 (25 tests), Non congelare. 9045-0792-120 (120 tests) Materiale fotosensibile. Clone: HM47 Attenzione: contiene azide Concentrazione: 5 µl (0,03 µg)/test (un test viene Manufacturer: eBioscience, Inc., 10255 Science definito come la quantità in grado di colorare Center Drive, San Diego, CA 92121, USA 1x10e6 cellule in 100 μl) Authorized Representative: Bender MedSystems Ospite/isotipo: IgG1 di topo, kappa GmbH, an eBioscience Company Campus Vienna Workshop HLDA: V Biocenter 2 A-1030 Vienna Austria Formulazione: Tampone acquoso, 0,09% di sodio azide; può contenere proteina carrier/stabilizzante. Uso previsto Descrizione L'anticorpo monoclonale HM47 coniugato con fluorocromo L'anticorpo monoclonale HM47 riconosce il dominio reagisce con l'antigene CD79a umano. Il CD79a può essere citoplasmatico del CD79a, chiamato anche mb-1. Il CD79a è rilevato in campioni biologici umani mediante tecniche una glicoproteina di membrana da 47 kDa che si unisce al immunologiche. CD79b con cui forma il recettore eterodimerico per i linfociti Principi del test B (BCR). Questo recettore è responsabile della segnalazione La citometria a flusso è uno strumento utile per la misurazione dei linfociti B e causa la loro attivazione, apoptosi o anergia.
    [Show full text]
  • Research Article Microarray Analysis of the Molecular Mechanism Involved in Parkinson’S Disease
    Hindawi Parkinson’s Disease Volume 2018, Article ID 1590465, 12 pages https://doi.org/10.1155/2018/1590465 Research Article Microarray Analysis of the Molecular Mechanism Involved in Parkinson’s Disease Cheng Tan, Xiaoyang Liu, and Jiajun Chen Department of Neurology, China-Japan Union Hospital of Jilin University, Changchun, Jilin 130033, China Correspondence should be addressed to Jiajun Chen; [email protected] Received 24 May 2017; Revised 21 August 2017; Accepted 18 October 2017; Published 1 March 2018 Academic Editor: Amnon Sintov Copyright © 2018 Cheng Tan et al. )is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Purpose. )is study aimed to investigate the underlying molecular mechanisms of Parkinson’s disease (PD) by bioinformatics. Methods. Using the microarray dataset GSE72267 from the Gene Expression Omnibus database, which included 40 blood samples from PD patients and 19 matched controls, differentially expressed genes (DEGs) were identified after data preprocessing, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Protein-protein interaction (PPI) network, microRNA- (miRNA-) target regulatory network, and transcription factor- (TF-) target regulatory networks were constructed. Results. Of 819 DEGs obtained, 359 were upregulated and 460 were downregulated. Two GO terms, “rRNA processing” and “cytoplasm,” and two KEGG pathways, “metabolic pathways” and “TNF signaling pathway,” played roles in PD development. Intercellular adhesion molecule 1 (ICAM1) was the hub node in the PPI network; hsa- miR-7-5p, hsa-miR-433-3p, and hsa-miR-133b participated in PD pathogenesis.
    [Show full text]
  • Analyzing the Phenotypic and Functional Complexity of Lymphocytes Using Cytof (Cytometry by Time-Of-Flight)
    Cellular & Molecular Immunology (2012) 9, 322–323 ß 2012 CSI and USTC. All rights reserved 1672-7681/12 $32.00 www.nature.com/cmi RESEARCH HIGHLIGHT Analyzing the phenotypic and functional complexity of lymphocytes using CyTOF (cytometry by time-of-flight) Guobing Chen and Nan-ping Weng Cellular & Molecular Immunology (2012) 9, 322–323; doi:10.1038/cmi.2012.16; published online 28 May 2012 he human immune system is equipped antigen peptide-MHC tetramers on human status of cytokines, cytotoxic granule T with vast numbers of lymphocytes pos- CD81 T cells. These parameters covered components and CD107, a degranulation sessing distinct antigen specificities. The nine functional attributes and distinguished marker. A shift in the proportions of enormous repertoire and heterogeneity of all reported CD81 T-cell subsets, such as cells with each phenotype was identified after lymphocytes ensures their ability to effec- naive, central memory, effector memory, in vitro stimulation with either PMA/ tively fight against external microbial inva- terminal effector, long-lived memory pre- ionomycin or anti-CD36anti-CD28. PMA/ ders, as well as internal aberrant cells. cursor effector cells and short-lived effector ionomycin induced a greater number of dis- However, the understanding of the specific cells. The authors used principal compon- tinct functional types of CD81 T cells than phenotypes and functions of lymphocytes is ent analysis (PCA) to analyze the large did anti-CD36anti-CD28. However, because hindered by the lack of tools that can be used CyTOF datasets and presented their results these studies were performed at the same to visualize this complexity. Fluorescent dye- either in two-dimensional PCA or in com- timepoint after stimulation, it is not clear based flow cytometry, which can simulta- bination with a protein structure program whether the observed difference in the num- 1 neously analyze 8–10 parameters of a single (PyMOL) as three-dimensional PCA.
    [Show full text]
  • A Possible Approach for Oral Drug Delivery of Nanoparticles
    COSMOS, Vol. 10, No. 1 (2014) 1–4 © World Scienti¯c Publishing Company DOI: 10.1142/S0219607714400035 A POSSIBLE APPROACH FOR ORAL DRUG DELIVERY OF NANOPARTICLES RABI'ATUL `ADAWIYAH BINTE MINHAT and THILO HAGEN Department of Biochemistry, Yong Loo Lin School of Medicine National University of Singapore, 117599 Singapore Received 12 February 2014 Revised 26 March 2014 Accepted 10 April 2014 Published 13 February 2015 Keywords: Oral drug delivery; nanoparticles; neonatal Fc receptor. Advances in biotechnology have led to numerous nanoparticles relative to the total atom number.2 discoveries and development of proteins and other These special properties are the reason why nano- macromolecules as pharmaceutical drugs.2 The particles have a wide variety of potential applica- production of macromolecules as therapeutics has tions in various ¯elds such as medicine, energy and improved treatment options in many areas in electronics industries. Thus, the development of an medicine. However, delivery of these macromolecule oral delivery system using nanoparticles is advan- COSMOS Downloaded from www.worldscientific.com drugs has mostly been restricted to parenteral tageous. If successful, the method could be combined methods of administration. The development of a with other techniques, such as a nanoparticle-based more convenient oral delivery system still faces many chemotherapy free of the debilitating side-e®ects,7,9 challenges. to further improve available treatment options. by NATIONAL UNIVERSITY OF SINGAPORE on 03/30/15. For personal use only. Many macromolecule drugs are vulnerable to the There have been several attempts to develop a pH variation and enzymatic degradation in the method for oral drug delivery using nanoparticles,2 gastrointestinal tract.2 One way to protect the drugs but these attempts have mostly had undesirable from degradation is by encapsulation.4 In particular, drawbacks.
    [Show full text]
  • A Role for the L-Selectin Adhesion System in Mediating Cytotrophoblast Emigration from the Placenta
    Developmental Biology 298 (2006) 107–117 www.elsevier.com/locate/ydbio A role for the L-selectin adhesion system in mediating cytotrophoblast emigration from the placenta ⁎ Akraporn Prakobphol a, Olga Genbacev a, Matthew Gormley a, Mirhan Kapidzic a, Susan J. Fisher a,b, a Department of Cell and Tissue Biology, University of California, San Francisco, CA 94143, USA b Departments of Anatomy and Pharmaceutical Chemistry, University of California, San Francisco, CA 94143, USA Received for publication 24 February 2006; revised 13 June 2006; accepted 13 June 2006 Available online 15 June 2006 Abstract Cytotrophoblast (CTB) aggregates that bridge the gap between the placenta and the uterus are suspended as cell columns in the intervillous space, where they experience significant amounts of shear stress generated by maternal blood flow. The proper formation of these structures is crucial to pregnancy outcome as they play a vital role in anchoring the embryo/fetus to the decidua. At the same time, they provide a route by which CTBs enter the uterine wall. The mechanism by which the integrity of the columns is maintained while allowing cell movement is unknown. Here, we present evidence that the interactions of L-selectin with its carbohydrate ligands, a specialized adhesion system that is activated by shear stress, play an important role. CTBs in cell columns, particularly near the distal ends, stained brightly for L-selectin and with the TRA-1-81 antibody, which recognizes carbohydrate epitopes that support binding of L-selectin chimeras in vitro. Function-perturbing antibodies that inhibited either receptor or ligand activity also inhibited formation of cell columns in vitro.
    [Show full text]
  • (CR1) and Crry, the Two Genetic Homologues of Human CR1 by Hector Molina,* Winnie Wong,~ Taroh Kinoshita,$ Carol Brenner,* Sharon Foley,* and V
    Disfin_ct Receptor and Regulatory Properties of Recombinant Mouse Complement Receptor 1 (CR1) and Crry, the Two Genetic Homologues of Human CR1 By Hector Molina,* Winnie Wong,~ Taroh Kinoshita,$ Carol Brenner,* Sharon Foley,* and V. Michael Holers* From the *Howard Hughes Medical Institute Laboratories and Department of Medicine, Division of Rheumatology, Washington University School of Medicine, St. Louis, Missouri 63110; the *BASF Bioresearch Corporation, Cambridge, Massachusetts 02139; and the SDepartraent of Immunoregulation, Research Institute for Microbial Diseases, Osaka University, Osaka 565, Japan Downloaded from http://rupress.org/jem/article-pdf/175/1/121/1267946/121.pdf by guest on 27 September 2021 Summary The relationship between the characterized mouse regulators of complement activation (RCA) genes and the 190-kD mouse complement receptor 1 (MCK1), 155-kD mouse complement receptor 2 (MCR2), and mouse p65 is unclear. One mouse RCA gene, designatedMCR2 (or Cr2), encodes alternatively spliced 21 and 15 short consensus repeat (SCR)-containing transcripts that crosshybridize with cDNAs of both human CR2 and CR1, or CR2 alone, respectively. A five SCR-containing transcript derived from a second unique gene, designated Crry, also crosshybridizes with human CR1. We have previously shown that the 155-kD MCR2 is encoded by the 15 SCR-containing transcript. To analyze the protein products of the other transcripts, which are considered the genetic homologues of human CRI, we have expressed the 21 and the 5 SCR-containing cDNAs in the human K562 erythroleukemia cell line. We demonstrate that cells expressing the 21 SCK transcript express the 190-kD MCR1 protein. These cells react with five unique rat anti-MCR1 monodonal antibodies, including the 8C12 antibody considered to be monospecific for MCK1.
    [Show full text]
  • Download Slides
    PEARLS OF LABORATORY MEDICINE Pearl Title: Immunoglobulin and T Cell Receptor Genetics Name of Presenter: Bing Melody Zhang Affiliation: Stanford University School of Medicine DOI: 10.15428/CCTC.2020.318055 © Clinical Chemistry Outline • Structure of Immunoglobulin (Ig) and TCR (T cell receptor) • TCR/Ig genetics • T/B cell development and TCR/Ig gene rearrangement • Molecular testing of TCR/Ig rearrangement • Clinical utility of molecular TCR/Ig rearrangement analysis in lymphoid malignancies 2 Immunoglobulin (Ig) and T Cell Receptor (TCR) 3 TCR/Ig Genetics • Each receptor chain is encoded in the germline genome as different gene segments (V, D, J). • During T/B lymphocyte development, the gene segments undergo random somatic DNA recombination (VDJ or VJ) to form a complete variable region sequence, known as gene rearrangement. • Non-templated insertion and deletion of random nucleotides at the junctional regions. • Somatic hypermutation: high rate of point mutations in V region of Ig genes (mature B cells). 4 IGH Gene Rearrangement V genes D genes J genes Constant region Germline V-N-D-N-J 5 B cell development and BCR/Ig gene rearrangement Large pre- Small pre- Immature B Mature B Stem cells Early pro-B Late pro-B cell cell B cell B cell cell cell IgH: germline D-J joining V-DJ joining VDJ rearranged VDJ rearranged VDJ rearranged VDJ rearranged IgK/L: germline germline germline germline V-J joining VJ rearranged VJ rearranged Order of gene rearrangements: IGH → IGK If IGK rearrangement fails → IGL gene rearrangement The Immune System,
    [Show full text]
  • TITLE: Immunoglobulin and T Cell Receptor Genetics PRESENTER: Bing Melody Zhang
    TITLE: Immunoglobulin and T Cell Receptor Genetics PRESENTER: Bing Melody Zhang Slide 1: Hello, my name is Bing Melody Zhang. I am an Assistant professor (Clinical) of Pathology at Stanford University. Welcome to this Pearl of Laboratory Medicine on “Immunoglobulin and T Cell Receptor Genetics.” Slide 2: Here is the outline of this talk. I will cover the structure and genetics of Ig and TCR, T/B cell development and TCR/Ig gene rearrangements, molecular testing and clinical utility of TCR/Ig rearrangement analysis. Slide 3: Let’s first review the structure of Ig and TCR. B and T lymphocytes are characterized by their antigen-specific receptors, which are the primary effector molecules of the adaptive immune system. For B cells, there are the membrane-bound form of the immunoglobulin as main component of the B cell receptor (BCR), and the soluble form of Ig secreted by plasma cells. Both forms of Ig have the identical Y- shaped structure, composed of 2 heavy and 2 light chains. © 2016 Clinical Chemistry Pearls of Laboratory Medicine Title On the other hand, T cell receptors are heterodimeric cell surface molecules responsible for recognizing antigens presented to T cells in context of MHC molecules. In the circulation, majority (95%) of T cells express alpha beta chains and a minority of T cells express gamma- delta chains, as part of the TCR complex. Gamma-delta T cells are dominant subtype of T cells found in epithelial tissues. Both TCR and Ig molecules consist of variable and constant regions. Each lymphocyte receptor has a unique antigen-specificity determined by the structure of antigen-binding site primarily from the variable amino acid sequences.
    [Show full text]
  • Cells of the Immune System and Innate Immunity
    Harvard-MIT Division of Health Sciences and Technology HST.176: Cellular and Molecular Immunology Course Director: Dr. Shiv Pillai Cells of the Immune System and Innate Immunity Recommended reading: Abbas et al., 4th edition, Chapters 2 and 12 Janeway and Travers, 5th edition, Chapters 1 and 10 The innate immune system is made up of molecules and cellular processes that defend the host within minutes and hours after exposure to a noxious stimulus. It is the only immune system in plants and invertebrates. Innate immune responses may initiate acute inflammation, which basically involves the accumulation and activation of phagocytes (mainly polymorphonuclear cells in man) and of vascular endothelium at sites of injury. In vertebrates, the innate and adaptive immune systems talk to each other and there is an important reciprocal interplay that operates between these systems. In these animals the cells and molecules of the innate immune system provide immediate protection and then set in motion the activation of the adaptive immune response. The adaptive immune response in turn “revs” up innate immune mechanisms of host defense. The innate immune system uses non-clonal “pattern recognition” receptors (PRRs) which cannot recognize host structures but which do recognize patterns found on microbes.These are sometimes called PAMPs for Pathogen Associated Molecular Patterns. Pathogens/ microbes The most common organisms that cause disease are viruses, bacteria, fungi, protozoa, and worms. A microbe (a small invader) or a worm that does not cause disease in most people may generally be classified as non-pathogenic. Such an organism will provoke an immune response in a normal immunocompetent host.
    [Show full text]
  • Tetraspanin 3, As a Novel Regulator of Intracellular Β1 Integrin Recycling, Promotes Non-Small Cell Lung Cancer Cell Proliferation
    Tetraspanin 3, as a Novel Regulator of Intracellular β1 Integrin Recycling, Promotes Non-Small Cell Lung Cancer Cell Proliferation Hongbo Su China Medical University Yitong Xu China Medical University Yao Zhang China Medical University Xizi Jiang China Medical University Qingfu Zhang China Medical University Hongjiu Ren China Medical University Zifang Zou China Medical University Xueshan Qiu ( [email protected] ) China Medical University https://orcid.org/0000-0001-9481-8249 Research Keywords: beta 1 integrin, tetraspanin 3, non-small cell lung carcinoma, Rab11a Posted Date: June 9th, 2021 DOI: https://doi.org/10.21203/rs.3.rs-572884/v1 License: This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full License Page 1/25 Abstract Background: The role of tetraspanins in cancer development has been widely reported. However, the expression and roles of tetraspanin 3 (TSPAN3) in solid tumors, including non-small cell lung cancer (NSCLC), have not yet been extensively investigated. In this study, we explored the role of TSPAN3 in NSCLC and its potential role in the tracking of integrin β1, which is highly expressed in this type of cancer. Methods: The expression of TSPAN3 in NSCLC tissue and NSCLC cell lines was evaluated, and the correlation between TSPAN3 expression and disease progression was assessed. Furthermore, loss- and gain-of-expression studies were conducted to determine the biological function of TSPAN3 both in vivo and vitro. The regulatory role of TSPAN3 in β1 integrin expression and intracellular recycling was studied through western blot, RT-PCR, and immunouorescence analyses. Results: TSPAN3 was found to be highly expressed in lung cancer cells and tissues.
    [Show full text]