Antigen experience relaxes the organisational structure of the T cell receptor repertoire
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Curated by eLife
Evaluation Summary:
This study will be of interest for the fields of T cell immunology and computational biology. It represents a careful descriptive study of the T cell repertoires of young and old mice, quantifying the relationships between naive, regulatory, effector and memory subsets. It represents a first step and would benefit from additional analyses and interpretations.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #3 agreed to share their name with the authors.)
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Abstract
The creation and evolution of the T cell receptor repertoire within an individual combines stochastic and deterministic processes. We systematically examine the structure of the repertoire in different T cell subsets in young, adult and LCMV infected mice, from the perspective of variable gene usage, nucleotide sequences and amino acid motifs. Young individuals share a high level of organization, especially in the frequency distribution of variable genes and amino acid motifs. In adult mice, this structure relaxes and is replaced by idiotypic evolution of the effector and regulatory repertoire. The repertoire of CD4+ regulatory T cells was more similar to naïve cells in young mice, but became more similar to effectors with age. Finally, we observed a dramatic restructuring of the repertoire following infection with LCMV. We hypothesize that the stochastic process of recombination and thymic selection initially impose a strong structure to the repertoire, which gradually relaxes following asynchronous responses to different antigens during life.
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Author Response:
Reviewer #1 (Public Review):
In this study, the authors present a detailed analysis of the T cell receptor repertoire in mice examining how the age, the cell differentiation status, the tissue compartment distinguishing between spleen and bone marrow, and antigen exposure influence its composition. Looking at amino acid motif distributions and sharing patterns of nucleotide sequences within the individual clones, and comparing them between the different cell compartments and groups, they aim at identifying the main axes of influence that shape the T cell receptor repertoire within these mice. They find some interesting differences, with e.g. repertoires from different functional compartments being more separated within young animals compared to older ones. However, given the complexity of the different aspects that …
Author Response:
Reviewer #1 (Public Review):
In this study, the authors present a detailed analysis of the T cell receptor repertoire in mice examining how the age, the cell differentiation status, the tissue compartment distinguishing between spleen and bone marrow, and antigen exposure influence its composition. Looking at amino acid motif distributions and sharing patterns of nucleotide sequences within the individual clones, and comparing them between the different cell compartments and groups, they aim at identifying the main axes of influence that shape the T cell receptor repertoire within these mice. They find some interesting differences, with e.g. repertoires from different functional compartments being more separated within young animals compared to older ones. However, given the complexity of the different aspects that are investigated and compared, it is sometimes difficult to follow the main conclusions from each of the presented analyses. In addition, the main conclusion shown in Figure 7 that repertoire evolution is influenced by cell migration, differentiation and age/infection seems to be partly well known, as also acknowledged by the authors. However, it would be really interesting if, based on their analyses, the authors could put a "weight" to each of these features, i.e. if aging has a larger effect on repertoire differences than tissue compartments. This is currently not easily seen from their analyses. If possible this would increase the value of this study going beyond a descriptive presentation of the results and support the mechanistic relationships hypothesized within the discussion.
We are grateful to the reviewer for these insightful comments. We have introduced a completely new analysis (new Figure 5 and two additional panels in Figure 6) which captures the quantitative shift in a two-dimensional space defined by two global statistical parameters of the repertoire (V and Triplet cosine similarity) as the repertoires move along the multidimensional trajectory illustrated in Figure 7 (old, now Figure 5A). This analysis framework is novel, at least to our knowledge. When combined with the existing analysis, this additional analysis reveals a clear hierarchy of impact of different immunological processes on repertoire diversification. Definition of this hierarchy is itself we believe novel. In addition, we observe a CD4+/CD8+ lineage dependent interaction between age and differentiation on the structure of the repertoire providing additional novelty. A completely new results and discussion session are attached to this figure.
Reviewer #2 (Public Review):
This is a careful observational study of a rich dataset, quantifying the relationships between naive, regulatory, effector and memory subsets. it is notable for its thorough approach to analysing TCR diversity by multiple levels of granularity, from V-beta usage to nucleotide level. However it is a little lacking in narrative and interpretation. As a result my impression is that it doesn't present any results that expand significantly on our existing understanding of T cell biology. As the authors note, shifts in diversity of T cell subsets with age are already well established and while multiple measures of diversity are explored, the results are broadly in agreement with each other. I wanted to be more enthusiastic about this study but it comes across as a tour de force in data exploration rather than something that sheds light on the forces shaping the structure and overlap of T cell repertoires.
We agree with this reviewer and the prior reviewer that the main impact of this study was not clearly explained. We provide a quantitative framework which can be used to track the movement of repertories along the multidimensional space illustrated in our Figure 7(old) and can define a hierarchy of impact of different immunological processes in driving diversification of the repertoire. As outlined in detail in the response to reviewer 1, we think this perspective on statistical features of TCR repertoires goes significantly beyond previous studies and provides a robust quantitative framework for analysing the relative influence of internal or external selective pressure on the T cell compartment.
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Evaluation Summary:
This study will be of interest for the fields of T cell immunology and computational biology. It represents a careful descriptive study of the T cell repertoires of young and old mice, quantifying the relationships between naive, regulatory, effector and memory subsets. It represents a first step and would benefit from additional analyses and interpretations.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #3 agreed to share their name with the authors.)
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Reviewer #1 (Public Review):
In this study, the authors present a detailed analysis of the T cell receptor repertoire in mice examining how the age, the cell differentiation status, the tissue compartment distinguishing between spleen and bone marrow, and antigen exposure influence its composition. Looking at amino acid motif distributions and sharing patterns of nucleotide sequences within the individual clones, and comparing them between the different cell compartments and groups, they aim at identifying the main axes of influence that shape the T cell receptor repertoire within these mice. They find some interesting differences, with e.g. repertoires from different functional compartments being more separated within young animals compared to older ones. However, given the complexity of the different aspects that are investigated and …
Reviewer #1 (Public Review):
In this study, the authors present a detailed analysis of the T cell receptor repertoire in mice examining how the age, the cell differentiation status, the tissue compartment distinguishing between spleen and bone marrow, and antigen exposure influence its composition. Looking at amino acid motif distributions and sharing patterns of nucleotide sequences within the individual clones, and comparing them between the different cell compartments and groups, they aim at identifying the main axes of influence that shape the T cell receptor repertoire within these mice. They find some interesting differences, with e.g. repertoires from different functional compartments being more separated within young animals compared to older ones. However, given the complexity of the different aspects that are investigated and compared, it is sometimes difficult to follow the main conclusions from each of the presented analyses. In addition, the main conclusion shown in Figure 7 that repertoire evolution is influenced by cell migration, differentiation and age/infection seems to be partly well known, as also acknowledged by the authors. However, it would be really interesting if, based on their analyses, the authors could put a "weight" to each of these features, i.e. if aging has a larger effect on repertoire differences than tissue compartments. This is currently not easily seen from their analyses. If possible this would increase the value of this study going beyond a descriptive presentation of the results and support the mechanistic relationships hypothesized within the discussion.
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Reviewer #2 (Public Review):
This is a careful observational study of a rich dataset, quantifying the relationships between naive, regulatory, effector and memory subsets. it is notable for its thorough approach to analysing TCR diversity by multiple levels of granularity, from V-beta usage to nucleotide level. However it is a little lacking in narrative and interpretation. As a result my impression is that it doesn't present any results that expand significantly on our existing understanding of T cell biology. As the authors note, shifts in diversity of T cell subsets with age are already well established and while multiple measures of diversity are explored, the results are broadly in agreement with each other. I wanted to be more enthusiastic about this study but it comes across as a tour de force in data exploration rather than …
Reviewer #2 (Public Review):
This is a careful observational study of a rich dataset, quantifying the relationships between naive, regulatory, effector and memory subsets. it is notable for its thorough approach to analysing TCR diversity by multiple levels of granularity, from V-beta usage to nucleotide level. However it is a little lacking in narrative and interpretation. As a result my impression is that it doesn't present any results that expand significantly on our existing understanding of T cell biology. As the authors note, shifts in diversity of T cell subsets with age are already well established and while multiple measures of diversity are explored, the results are broadly in agreement with each other. I wanted to be more enthusiastic about this study but it comes across as a tour de force in data exploration rather than something that sheds light on the forces shaping the structure and overlap of T cell repertoires.
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Reviewer #3 (Public Review):
The adaptive immune system employs an incredibly diverse set of non-germline-encoded receptors for specific defense. Despite this diversity the system responds robustly to infections across individuals, and there is even a certain degree of stereotypy in the targets of response. A key question in the field is how diversity and robustness can co-exist and how both are sculpted by the stochastic processes shaping an individual's immune repertoire. Here, Mark et al. address this question in a tour de force combining extensive repertoire sequencing and novel computational analysis techniques. A major strength of the manuscript is the detailed dataset that underlies the findings: The authors have used sequencing to map repertoire organization across multiple phenotypic and spatial compartments in mice, including …
Reviewer #3 (Public Review):
The adaptive immune system employs an incredibly diverse set of non-germline-encoded receptors for specific defense. Despite this diversity the system responds robustly to infections across individuals, and there is even a certain degree of stereotypy in the targets of response. A key question in the field is how diversity and robustness can co-exist and how both are sculpted by the stochastic processes shaping an individual's immune repertoire. Here, Mark et al. address this question in a tour de force combining extensive repertoire sequencing and novel computational analysis techniques. A major strength of the manuscript is the detailed dataset that underlies the findings: The authors have used sequencing to map repertoire organization across multiple phenotypic and spatial compartments in mice, including following an external challenge (LCMV infection) and in two age groups. Among the notable findings are the following: First, a number of high-level statistical observables (V gene/amino acid motif usage) show little variability in the naive compartment of young mice, which increases both with age and in effector/memory repertoires. Second, there are statistical features that discriminate between different compartments, most clearly for the CD4/CD8 subsets but also between phenotypic subsets. Importantly, the paper skillfully connects the somewhat abstact study of repertoire organization to concrete questions about the underlying immunobiology, thus demonstrating the usefulness of the architectural viewpoint. For instance, the analyses show that the Treg compartment transitions from a naive-like to an effector-like composition with age, presumably reflecting the natural vs. induced Treg shift.
Overall the claims are well-supported by the presented analyses, but the evidence could be further strengthened by a more detailed methods section and additional robustness checks with regards to sampling variations.
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