Bcl6 is the nexus transcription factor of T follicular helper cell (TFH) differentiation via a set of repressor-of-repressor gene circuits

Abstract

Abstract T follicular helper (TFH) cells are a distinct type of CD4 T cells that are essential for most antibody and B lymphocyte responses. TFH regulation and dysregulation is involved in a range of diseases. Bcl6 is the lineage defining transcription factor of TFH cells and its activity is essential for TFH cell differentiation and function. However, how Bcl6 controls TFH biology has largely remained unclear, at least in part due to intrinsic challenges of connecting repressors to gene upregulation in complex cell types with multiple possible differentiation fates. Multiple competing models were tested here by a series of experimental approaches. We ruled out the model that TFH is a default pathway of CD4 T cell by utilizing Bcl6 and Blimp1 double deficient mice in acute virus infection and protein immunization. We showed that Bcl6 has a strong negative autoregulatory loop in TFH cells, by generating mice containing a deletion mutation of an 8-nucleotide Bcl6 binding site in the Bcl6 promoter. To determine putative Bcl6 target TFs, we developed an integrated analytic approach using RNA-seq, BCL6 ChIP-seq, and ATAC-seq data and Bcl6 and Blimp1 double KO mice. Using this approach, Runx2, Runx3, and Klf2 were identified as Bcl6-targeted repressors (Bcl6-r TFs) that regulate important TFH genes. Taken together, Bcl6 controls pleiotropic attributes of TFH differentiation and function (migration, costimulation, inhibitory receptors, and cytokines) via multiple repressor-of-repressor gene circuits.