T cell help transiently unlocks a high plasticity state in germinal center B cells during the humoral immune response [iPSC_RNA-seq]

During cellular differentiation, phenotypic plasticity is gradually lost due to the strengthening of epigenetic barriers, although it may be re-acquired upon disease or injury. Here, we report an unprecedented physiological gain of epigenetic plasticity among mature B cells during the humoral immune response. Acquisition of plasticity was strictly dependent on T follicular helper (TFH) cells and restricted to germinal center (GC) but not activated B cells, indicating both non- and cell-autonomous contributions to this phenotype. Importantly, GC plasticity was not dependent on proliferation and could not be solely explained by the activation of MYC programs. Instead, GC plasticity was associated with the reciprocal weakening of B cell identity and the de- repression of progenitor and stem cell-like programs induced by NF?B and other signals, downstream of TFH signaling. Therefore, physiological acquisition of GC B cell plasticity is tightly controlled by the affinity maturation positive selection checkpoint. Notably, Histone 1 loss of function, which results in aberrant stem cell gene reactivation through chromatin decompaction, allowed GC B cells to bypass this gatekeeping mechanism. Finally, patients with Diffuse LargeB Cell Lymphoma (DLBCL) that enriched for GC plasticity signatures had significantly worse outcomes, implicating a role for this mechanism in immune regulation and fitness and potential relevance to lymphomagenesis. Overall design: Splenic B cells from mice (males) immunized with SRBC for 9 days were pre-enriched for B220pos cells by negative magnetic enrichment and stained for germinal center B cell markers (B220+ CD38- Fas+), including the Myc-GFP endogeneous reported for isolation of TFH-activated GC B cells by flow sorting. Bulk GC or TFH-activated GC B cells were plated on Mouse Embryonic Fibroblasts (MEF) for 6 days with cytokines for somatic cell reprogramming to induced Pluripotent Stem Cells (iPSC) upon treatment with Doxycyline (Dox). After 10 days of reprogramming, the iPSC colonies emerging were culture in abscence of Dox for 4 additional days to select for truly transgene-independent and pluripotent iPSCs. 3 individual colonies (clones) were manually picked and isolated from each Bulk GC or Myc-GFP GC-derived iPSCs. Cells were passaged once on gelatin only and pre-plated for an additional 45 min to remove the remaining MEF. Collected cells were processed for RNA extraction using the RNAeasy Qiagen kit according to the provider specifications. RNAs sent to Novogene Bioinformatics Technology Co. Ltd. for QC and Libraries preparation (non-stranded Poly A) and clustered on an Illumina NovaSeq X Plus Series (Pair End 150)