Dynamics of Individual T Cell Repertoires: From Cord Blood to Centenarians Olga V. Britanova, Mikhail Shugay, Ekaterina M. Merzlyak, Dmitriy B. Staroverov, Ekaterina V. Putintseva, Maria A. This information is current as Turchaninova, Ilgar Z. Mamedov, Mikhail V. Pogorelyy, of September 27, 2021. Dmitriy A. Bolotin, Mark Izraelson, Alexey N. Davydov, Evgeny S. Egorov, Sofya A. Kasatskaya, Denis V. Rebrikov, Sergey Lukyanov and Dmitriy M. Chudakov J Immunol published online 13 May 2016 Downloaded from http://www.jimmunol.org/content/early/2016/05/12/jimmun ol.1600005 Supplementary http://www.jimmunol.org/content/suppl/2016/05/12/jimmunol.160000 http://www.jimmunol.org/ Material 5.DCSupplemental Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists by guest on September 27, 2021 • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2016 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. Published May 13, 2016, doi:10.4049/jimmunol.1600005 The Journal of Immunology Dynamics of Individual T Cell Repertoires: From Cord Blood to Centenarians Olga V. Britanova,*,†,‡,1 Mikhail Shugay,*,†,‡,1 Ekaterina M. Merzlyak,*,1 Dmitriy B. Staroverov,*,1 Ekaterina V. Putintseva,* Maria A. Turchaninova,*,†,‡ Ilgar Z. Mamedov,*,‡ Mikhail V. Pogorelyy,* Dmitriy A. Bolotin,*,†,‡ Mark Izraelson,*,‡ Alexey N. Davydov,‡ Evgeny S. Egorov,*,†,‡ Sofya A. Kasatskaya,* Denis V. Rebrikov,†,x Sergey Lukyanov,*,† and Dmitriy M. Chudakov*,†,‡ The diversity, architecture, and dynamics of the TCR repertoire largely determine our ability to effectively withstand infections and malignancies with minimal mistargeting of immune responses. In this study, we have employed deep TCRb repertoire sequencing with normalization based on unique molecular identifiers to explore the long-term dynamics of T cell immunity. We demonstrate Downloaded from remarkable stability of repertoire, where approximately half of all T cells in peripheral blood are represented by clones that persist and generally preserve their frequencies for 3 y. We further characterize the extremes of lifelong TCR repertoire evolution, analyzing samples ranging from umbilical cord blood to centenarian peripheral blood. We show that the fetal TCR repertoire, albeit structurally maintained within regulated borders due to the lower numbers of randomly added nucleotides, is not limited with respect to observed functional diversity. We reveal decreased efficiency of nonsense-mediated mRNA decay in umbilical cord blood, which may reflect specific regulatory mechanisms in development. Furthermore, we demonstrate that human TCR rep- http://www.jimmunol.org/ ertoires are functionally more similar at birth but diverge during life, and we track the lifelong behavior of CMV- and EBV- specific T cell clonotypes. Finally, we reveal gender differences in dynamics of TCR diversity constriction, which come to naught in the oldest age. Based on our data, we propose a more general explanation for the previous observations on the relationships between longevity and immunity. The Journal of Immunology, 2016, 196: 000–000. he native human TCR repertoire changes throughout life specificities that is available to respond to new challenges and to based on Ag experience, exhaustion, and nonuniformity sustain a balanced network of regulatory interactions (1, 7–12). T of T cell proliferation at the periphery, as well as on the The potential for revealing a detailed view of individual TCR by guest on September 27, 2021 aging-associated evolution and involution of the thymus (1–6). At repertoires, which opened with development of high-throughput any given moment, our accumulated population of expanded clonal sequencing techniques (13–16), has attracted many laboratories to T cells represents our up-to-date Ag experience and determines the this presently superficially explored field. Multiple studies have current state of our adaptive immune defenses, whereas the remaining been performed in recent years to characterize TCR repertoires diversity of the naive T cell population represents the reservoir of new either in general (13–22) or at the level of functional T cell subsets (17, 18, 23–33). However, these new opportunities have also brought new hidden pitfalls, which have not always been predicted, *Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry of the Russian Acad- emy of Sciences, Moscow 117997, Russia; †Pirogov Russian National Research recognized, and proportionally addressed in the works published to Medical University, Moscow 117997, Russia; ‡Central European Institute of Tech- date. As long as the analyzed material is of sufficient quality and x nology, Masaryk University, 625 00 Brno, Czech Republic; and Vavilov Institute of quantity, shallow profiling at the scale of up to several thousands of General Genetics of the Russian Academy of Sciences, Moscow 119991, Russia 1 expanded clonotypes can usually be reliably interpreted. At the O.V.B., M.S., E.M.M., and D.B.S. contributed equally to this work. same time, deep and reliable profiling of hundreds of thousands and ORCIDs: 0000-0001-7826-7942 (M.S.); 0000-0001-6789-9442 (D.B.S.); 0000-0003- millions of analyzed T cells and sequencing reads remains chal- 4421-3844 (E.V.P.); 0000-0002-8484-6067 (D.A.B.); 0000-0002-6495-7173 (E.S.E.); 0000-0002-5522-7443 (S.A.K.); 0000-0002-1884-1807 (D.V.R.); 0000-0003-0430- lenging. This is compounded by biased amplification and accu- 790X (D.M.C.). mulation of PCR and sequencing errors (34–39), which must be Received for publication January 6, 2016. Accepted for publication April 16, 2016. distinguished from the true highly homologous TCR variants This work was supported by Russian Science Foundation Project 16-15-00149. M.S. originating from convergent recombination events (18, 19, 40). and E.V.P. are supported by individual fellowship Russian Foundation for Basic Potential cross-sample contamination and sequencing artifacts can Research Grants 16-34-60179 and 16-34-60178. further hamper accurate comparison of TCR repertoires in different The raw sequencing data in this article have been submitted to the National Center for Biotechnology Information Sequence Read Archive database (https://www.ncbi. samples. As we have accumulated more experience with deep TCR nlm.nih.gov/Traces/sra/) under accession number PRJNA316572. repertoire profiling, these limitations have become clearer, along Address correspondence and reprint requests to Dr. Dmitriy M. Chudakov, Shemyakin– with the possible solutions for obtaining more quantitative data on Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, clonotype diversity, minimizing the generation of artificial and loss Miklukho-Maklaya 16/10, Moscow 117997, Russia. E-mail address: [email protected] of real receptor diversity (35, 37–39, 41, 42). The online version of this article contains supplemental material. In this study, we employed our present molecular and software Abbreviations used in this article: MIG, molecular identifier group; NMD, nonsense- solutions to track both deeply and quantitatively the individual mediated decay; UCB, umbilical cord blood. dynamics and stability of T cell clones during 3 y. We also reveal Copyright Ó 2016 by The American Association of Immunologists, Inc. 0022-1767/16/$30.00 characteristic features of TCR repertoire at the polar extremes of www.jimmunol.org/cgi/doi/10.4049/jimmunol.1600005 2 DYNAMICS OF INDIVIDUAL T CELL REPERTOIRES the human life, including umbilical cord blood (UCB) and peripheral described previously (5) and analyzed via Illumina HiSeq 2500 blood samples obtained from centenarians, and we describe the paired-end 100 + 100 nt sequencing. In our downstream bio- general dynamics and extent of changes in the structure of the TCR informatics analysis, we grouped sequencing reads labeled with repertoire throughout life. Furthermore, we reveal gender differ- the same unique cDNA identifier to the MIG. Each nucleotide ences in adaptive immunity aging and link this finding to previous within the TCRb CDR3 sequence in each MIG was identified data on population studies. according to the cumulative quality in the assembled sequencing reads. This approach improves sequencing quality and eliminates Materials and Methods most PCR and sequencing errors according to the Safe-seqS logic Sample collection described by Kinde et al. (44). Because MIG-based analysis counts initial cDNA events, it also eliminates amplification and This study was approved by the local ethics committee and conducted in ac- sequencing biases and artifacts and thus significantly improves cordance with the Declaration of Helsinki. All donors were informed of the final use of their blood and signed an informed consent document. The cohort in- quantitative power in repertoire data analyses (5, 39, 45–48). cluded 65 healthy individuals aged 6–103 y (see Table II). We excluded indi- Importantly,