European Network linking informatics and genomics of helper T cells

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Unveiling the regulation of germinal centre responses and antibody production by follicular T cells

Publication . Kumar, Saumya; Graça, Luís Ricardo Simões da Silva; Carvalho, Margarida Henriques da Gama

Germinal Centres (GCs) are the sites in secondary lymphoid tissues specialized in the selection of the effective antibody response against invading pathogens. Within GCs, B cells undergo class switch recombination (CSR) and somatic hyper-mutation (SHM) followed by the selection of high-affinity clones in a T-cell-dependent manner. The GC T cells specialized in providing help to B cells, called T follicular helper (Tfh) cells, control and coordinate the most adequate antibody production. Another subset of follicular T cells, known as T follicular regulatory (Tfr) cells, have an important function in maintaining immune homeostasis by suppressing the generation of autoantibodies. These contrasting roles of Tfh and Tfr cells are essential for efficient GC reactions, by promoting effective humoral responses against pathogens while protecting the host from autoimmunity. It has been postulated that, in addition to the selection of affinity-matured B cell clones, Tfh cells may also provide instructions to drive appropriate isotype selection. However, the phenotype of these distinct specialized Tfh cells has remained poorly characterized. The first part of this work focused on elucidating the characteristics of specialized Tfh populations that emerge under type-1 and type-2 immune responses. To attain this, we generated whole-genome bulk and single-cell transcriptomics datasets of Tfh cells by collecting nearly homogenous Tfh populations using adjuvant immunizations biased towards type 1 or type 2 responses. We derived a transcriptional signature of Tfh1 and Tfh2 subsets using a machine learning approach and validated this signature by correctly classifying public datasets of Tfh cells from LCMV and Helminth’s infections.Further, single datasets shed light on the heterogeneity of Tfh1 and Tfh2 cells under different immunizations. These results also showed potential differences in the spatial distribution of Tfh subsets, thereby highlighting differences in Tfh help to B cells under the two types of the immune response. The molecular mechanisms underlying the function of human Tfh and Tfr cells remains poorly addressed, in part due to difficult access to lymphoid tissues where these cells reside. Both Tfr and Tfh cells share key features. However, these cells show a heterogeneous profile when examined in different tissues. The second part of this work focused on establishing a relationship of Tfh and Tfr cells across different lymphoid tissues in humans. We performed index-sort-based single-cell RNA-seq analysis of human Tfh and Tfr cells from blood, lymph nodes, and tonsils. We first examined Tfh cells from the three tissues and were able to re-create their maturation trajectory using pseudo-time ordering of these cells based on their transcriptome. When we examined the maturation of Tfr cells using RNA-velocity we found that Tfr cells follow a bifurcated trajectory, originating from Treg cells and differentiating into either immature blood Tfr cells, or mature Tfr cells that seed GCs in lymphoid tissues. Flow cytometry analysis of key molecular markers further confirmed these results. In conclusion, this work took advantage of transcriptomics datasets and computational methods to identify unique characteristics of Tfh subsets as well as to establish the relationship among human Tfh and Tfr cells across different tissues.

2022-02Tese de doutoramento

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Entidade financiadora

European Commission

Programa de financiamento

H2020

Número da atribuição

675395