Project description:This experiment was performed to identify somatic hypermutations in germinal center B cells of a mouse expressing the inferred germline of IOMA BCR after and a 5 step sequential immunization regimen.
Project description:Productive B cell responses are critical to protect a host from infection. The spleen and lymph nodes are populated by resting follicular B cells, which can enter germinal centers upon antigen encounter. Once in the germinal center, B cells migrate between the dark and light zones, where they undergo somatic hypermutation and selection, respectively. While germinal center B cells have been studied, an intense molecular understanding of these cells/subsets (and the differences between them) is lacking.
Project description:Germinal center B cells were isolated from human tonsil tissue and crosslinked. ChIP was performed on two distinct pools of germinal center cells, each obtained from 3-5 donors. The experiment includes two biological replicates (germinal center cell pools from different donors). ChIP was performed on both pools and subject to library preparation and sequencing. Input DNA was sequenced for both pools.
Project description:Given the tumor suppressing function of miR-15a/16-1 cluster, we studied its role in the germinal center B-cells that give rise to most lymphoid malignancies.
Project description:B cells positive for Ig kappa and Ig lambda are observed by flow cytometry in one fourth of patients with Systemic Lupus Erythematosus (SLE). Single cell Ig VDJ sequencing (10X Genomics) reveals that kappa/lambda B cells are at the same frequency (about 1.5%) in these SLE patients as in healthy controls. Cells observed by flow cytometry are instead decorated with VH4-34 IgM (kappa or lambda) autoantibodies that are present in some but not all SLE patients.
Project description:Germinal center B cells were isolated from human tonsil tissue and crosslinked. ChIP was performed on two distinct pools of germinal center cells, each obtained from 3-5 donors.
Project description:We report the gene expression profiles of germinal center B cells obtained by FACS analyses of normal human lymph nodes. We used FACS analyses to isolate germinal center B cells from 3 different individuals with one individual sampled twice. The overall goal was simply to identify genes that are highly expressed or silent in these purified cell populations.
Project description:The Germinal center is a dynamic microenvironment wherein B cells expressing high affinity antibody variants produced by hypermutation are selected for clonal expansion by limiting numbers of T follicular helper cells. Although a great deal is known about the mechanisms that control B cell selection in the germinal center, far less is understood about the clonal behavior of the T follicular helper cells that regulate this process. Here we report on the dynamic behavior of clones of T follicular helper cells during the germinal center reaction. We find that like germinal center B cells, T follicular helper cells undergo antigen dependent selection during the germinal center reaction resulting in differential proliferative expansion and contraction. Increasing the amount of antigen presented in the germinal center leads to increased T follicular cell division. Competition between T follicular helper cell clones is mediated by T cell receptor affinity for peptide-MHC ligand. Higher affinity T cells expanding preferentially in the germinal center show increased expression of genes downstream of the T cell receptor, genes required for metabolic reprogramming, cell division and cytokine production. These dynamic changes lead to dramatic remodeling of the functional T follicular cell repertoire during the germinal center reaction.
Project description:The chemokines CXCL13 and CXCL12 are reported to be important for the germinal center reaction. Since CXCL12-deficient mice are embryonically lethal, here we took advantage of the Cxcl13-Cre/TdTomato mouse models to genetically ablate CXCL12 from B cell-interacting reticular cells and examine the molecular consequence on germinal center B cells. Spatial segregation of follicular dendritic cells, germinal center B cells and follicular helper T cells is impaired in Cxcl13-Cre/TdTomato Cxcl12fl/fl mice. Single cell transcriptomic analysis revealed that all germinal center B cell subsets (corresponding to distinct stages of the germinal center response) are present in draining lymph nodes of immunized CXCL12-conditionally deficient mice. While most transcriptional regulators of the germinal center response are unperturbed by the genetic perturbation of CXCL12, Bach2 levels were elevated in germinal center B cells from lymph nodes of Cxcl12fl/fl mice. Moreover, single cell B cell receptor sequencing revealed that germinal center B cells in Cxcl13-Cre/TdTomato Cxcl12fl/fl mice harbour a lower mutational burden when compared to germinal center B cells isolated from immunized control mice. Gene expression profiles were validated by flow cytometry and suggest that the provision of CXCL12 by reticular cells governs efficient germinal center responses.