The Igκ3′ enhancer is activated by gradients of chromatin accessibility and protein association

The Igkappa locus is recombined following initiation of a signaling cascade during the early pre-B stage of B cell development. The Ig kappa3' enhancer plays an important role in normal B cell development by regulating kappa locus activation. Quantitative analyses of kappa3' enhancer chromatin structure by restriction endonuclease accessibility and protein association by chromatin immunoprecipitation in a developmental series of primary murine B cells and murine B cell lines demonstrate that the enhancer is activated progressively through multiple steps as cells mature. Moderate kappa3' chromatin accessibility and low levels of protein association in pro-B cells are increased substantially as the cells progress from pro- to pre-B, then eventually mature B cell stages. Chromatin immunoprecipitation assays suggest transcriptional regulators of the kappa3' enhancer, specifically PU.1 and IFN regulatory factor-4, exploit enhanced accessibility by increasing association as cells mature. Characterization of histone acetylation patterns at the kappa3' enhancer and experimental inhibition of histone deacetylation suggest changes therein may determine changes in enzyme and transcription factor accessibility. This analysis demonstrates kappa activation is a multistep process initiated in early B cell precursors before Igmu recombination and finalized only after the pre-B cell stage.