Spi-C and PU.1 counterregulateRag1andIgκ transcription to effect immunoglobulin kappa recombination in small pre-B cells

Abstract

AbstractB cell development requires the ordered rearrangement ofIggenes encoding H and L chain proteins that assemble into BCRs or Abs capable of recognizing specific Ags.Igκ rearrangement is promoted by chromatin accessibility and by relative abundance of RAG1/2 proteins. Expression of the E26-transformation-specific (ETS) transcription factor Spi-C is activated in response to double-stranded DNA breaks (DSBs) in small pre-B cells to negatively regulate pre-BCR signaling andIgκ rearrangement. However, it is not clear if Spi-C regulatesIgκ rearrangement through transcription or by controlling RAG expression. In this study, we investigated the mechanism of Spi-C negative regulation ofIgκ light chain rearrangement. Using an inducible expression system in a pre-B cell line, we found that Spi-C negatively regulatedIgκ rearrangement,Igκ transcript levels, andRag1transcript levels. We found thatIgκ andRag1transcript levels were increased in small pre-B cells fromSpic-/-mice. In contrast,Igκ andRag1transcript levels were activated by PU.1 and were decreased in small pre-B cells from PU.1-deficient mice. Using chromatin immunoprecipitation analysis, we identified an interaction site for PU.1 and Spi-C located in theRag1promoter region. These results suggest that Spi-C and PU.1 counterregulateIgκ transcription andRag1transcription to effectIgκ recombination in small pre-B cells.