Human germinal center transcriptional programs are de-synchronized in B cell lymphoma

Milpied, Pierre; Cervera-Marzal, Iñaki; Mollichella, Marie-Laure; Tesson, Bruno; Brisou, Gabriel; Traverse-Gléhen, Alexandra; Salles, Gilles; Spinelli, Lionel; Nadel, Bertrand

doi:10.1038/s41590-018-0181-4

Cited by 126 publications

(159 citation statements)

References 56 publications

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“…During affinity maturation and selection in the GC, B cells cycle between physically distinct light and dark zones. While we clearly identified LZ and DZ B cell populations in our scRNA-seq dataset, we also found that many GC B cells existed in a continuum between these two states (Figure 4A-B) similar to previous reports (Milpied et al, 2018), with the exception of the FCRL2/3 high and prePB clusters which existed as transcriptionally distinct states (Figures 2H and 4C). In addition to unique gene expression patterns, these two sub-populations of GC B cells also exhibit unique patterns of class switching, with prePB B cells in the GC more likely to express class-switched isotypes and FCRL2/3 high GC B cells more likely to retain expression of IgM and IgD (Figures 2I and 4D).…”

Section: Resultssupporting

confidence: 90%

Antibody repertoire and gene expression dynamics of diverse human B cell states during affinity maturation

King

Orban

Riches

et al. 2020

Preprint

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15In response to antigen challenge, B cells clonally expand, undergo selection and differentiate to produce 16 mature B cell subsets and high affinity antibodies. However, the interplay between dynamic B cell states 17 and their antibody-based selection is challenging to decipher in primary human tissue. We have applied 18 an integrated analysis of bulk and single-cell antibody repertoires paired with single-cell transcriptomics 19 of human B cells undergoing affinity maturation. We define unique gene expression and antibody 20 repertoires of known and novel B cell states, including a pre-germinal centre state primed to undergo 21 class switch recombination. We dissect antibody class-dependent gene expression of germinal centre 22 and memory B cells to find that class switching prior to germinal centre entry dictates the capacity of B 23 cells to undergo antibody-based selection and differentiate. Together, our analyses provide 24 unprecedented resolution into the gene expression and selection dynamics that shape B cell-mediated 25 immunity. 26 27 121 122 157 sampling of IgH sequences from bulk repertoires to enhance genotype correction, identification of 158

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Section: Resultssupporting

confidence: 90%

Antibody repertoire and gene expression dynamics of diverse human B cell states during affinity maturation

King

Orban

Riches

et al. 2020

Preprint

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“…The expression levels of surface protein markers recorded through index sorting were consistent with the gating strategy of Mem B cells (CD20 + CD38 lo CD10 -CD27 + ), GC B cells (CD20 + CD38 + CD10 + ) and PB/PCs (CD38 hi CD27 hi ) (Figure 4A Figure 4B), and identified the top marker genes for each cell subset ( Figure 4C). Those analyses were consistent with previous single-cell qPCR analyses 17 and bulk microarray analyses of human B cell subsets 27,28 .…”

Section: Fb5p-seq Analysis Of Human Tonsil B Cell Subsetssupporting

confidence: 90%

“…The progeny of a single T or B cell can thus be accurately identified through identical (for TCR) or very similar (for BCR) V-J junctional sequences in their TCR or BCR chain genes, respectively. Integrating single-cell immunoglobulin heavy chain (IGH) sequencing with lowthroughput gene expression analysis by single-cell qPCR already revealed important features of memory B cell diversification 16 and B cell lymphoma evolution 17 . Methods which enable parallel analysis of repertoire sequence and whole transcriptome gene expression in single B or T cells are required to deeply investigate multiple aspects of lymphocyte biology and malignancy.…”

Section: Introductionmentioning

confidence: 99%

FB5P-seq: FACS-based 5-prime end single-cell RNAseq for integrative analysis of transcriptome and antigen receptor repertoire in B and T cells

Attaf

Cervera-Marzal

Dong

et al. 2019

Preprint

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Single-cell RNA sequencing (scRNA-seq) allows the identification, characterization, and quantification of cell types in a tissue. When focused on B and T cells of the adaptive immune system, scRNA-seq carries the potential to track the clonal lineage of each analyzed cell through the unique rearranged sequence of its antigen receptor (BCR or TCR, respectively), and link it to the functional state inferred from transcriptome analysis. Here we introduce FB5Pseq, a FACS-based 5'-end scRNA-seq method for cost-effective integrative analysis of transcriptome and paired BCR or TCR repertoire in phenotypically defined B and T cell subsets. We describe in details the experimental workflow and provide a robust bioinformatics pipeline for computing gene count matrices and reconstructing repertoire sequences from FB5P-seq data. We further present two applications of FB5P-seq for the analysis of human tonsil B cell subsets and peripheral blood antigen-specific CD4 T cells. We believe our novel integrative scRNA-seq method will be a valuable option to study rare adaptive immune cell subsets in immunology research.B cell subsets and antigen-specific peripheral blood T cells, highlighting the relevance and performance of our cost-effective and scalable technology.

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“…3C). In the tumor biopsy from a freshly diagnosed follicular lymphoma (FL) (36), n = 13 γδ T lymphocytes of 472 TILs (3.3%) were detected ( Fig. 3D and SI Appendix, Fig.…”

Section: A Gene Signature To Detect Human γδ T Lymphocytes In Complexmentioning

confidence: 99%

Single-cell RNA sequencing unveils the shared and the distinct cytotoxic hallmarks of human TCRVδ1 and TCRVδ2 γδ T lymphocytes

Pizzolato

Kaminski

Tosolini

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

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γδ T lymphocytes represent ∼1% of human peripheral blood mononuclear cells and even more cells in most tissues of vertebrates. Although they have important anticancer functions, most current single-cell RNA sequencing (scRNA-seq) studies do not identify γδ T lymphocytes because their transcriptomes at the single-cell level are unknown. Here we show that high-resolution clustering of large scRNA-seq datasets and a combination of gene signatures allow the specific detection of human γδ T lymphocytes and identification of their T cell receptor (TCR)Vδ1 and TCRVδ2 subsets in large datasets from complex cell mixtures. In t-distributed stochastic neighbor embedding plots from blood and tumor samples, the few γδ T lymphocytes appear collectively embedded between cytotoxic CD8 T and NK cells. Their TCRVδ1 and TCRVδ2 subsets form close yet distinct subclusters, respectively neighboring NK and CD8 T cells because of expression of shared and distinct cytotoxic maturation genes. Similar pseudotime maturation trajectories of TCRVδ1 and TCRVδ2 γδ T lymphocytes were discovered, unveiling in both subsets an unattended pool of terminally differentiated effector memory cells with preserved proliferative capacity, a finding confirmed by in vitro proliferation assays. Overall, the single-cell transcriptomes of thousands of individual γδ T lymphocytes from different CMV + and CMV − donors reflect cytotoxic maturation stages driven by the immunological history of donors. This landmark study establishes the rationale for identification, subtyping, and deep characterization of human γδ T lymphocytes in further scRNA-seq studies of complex tissues in physiological and disease conditions. γδ T lymphocyte | transcriptome | single-cell RNA-sequencing | human immunology | cancer S ingle-cell level mRNA-sequencing (scRNA-seq) of heterogeneous cell populations has become the reference tool for establishing cellular lineages and composition of tissues from the human body (1). In addition, the development of novel and open-source computational tools for processing scRNA-seq datasets enables delineation of both broad and subtle differences in rare cell subsets present in complex mixtures, such as peripheral blood mononuclear cells (PBMC) (2, 3). Such developments are expected to build more knowledge about the cellular composition and states composing tumors. Recent scRNA-seq analyses in melanoma and colorectal cancer evidenced the various tumor microenvironments and exhaustion patterns of the tumor-infiltrating lymphocytes (4). Most of such studies currently focus on the cytotoxic CD8 T lymphocytes, which represent the main target of immune checkpoint blockade therapies, and are readily detected by their scRNA-seq profile. Other subsets of cytolytic T cells are also critical in this therapeutic perspective; however, currently they have never been characterized by scRNA-seq and are therefore missing from all current tumor microenvironments mapped by these technologies.Human γδ T lymphocytes represent a peculiar lymphoid cell subset displaying hall...

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Human germinal center transcriptional programs are de-synchronized in B cell lymphoma

Cited by 126 publications

References 56 publications

Antibody repertoire and gene expression dynamics of diverse human B cell states during affinity maturation

Antibody repertoire and gene expression dynamics of diverse human B cell states during affinity maturation

FB5P-seq: FACS-based 5-prime end single-cell RNAseq for integrative analysis of transcriptome and antigen receptor repertoire in B and T cells

Single-cell RNA sequencing unveils the shared and the distinct cytotoxic hallmarks of human TCRVδ1 and TCRVδ2 γδ T lymphocytes

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