Bruton's Tyrosine Kinase Regulates Immunoglobulin Promoter Activation in Association with the Transcription Factor Bright

Abstract

Bright (B-cell regulator of immunoglobulin heavy chain transcription) binding to immunoglobulin heavy chain loci after B-cell activation is associated with increased heavy chain transcription. Our earlier reports demonstrated that Bright coimmunoprecipitates with Bruton's tyrosine kinase (Btk) and that these proteins associate in a DNA-binding complex in primary B cells. B cells from immunodeficient mice with a mutation in Btk failed to produce stable Bright DNA-binding complexes. In order to determine if Btk is important for Bright function, a transcription activation assay was established and analyzed using real-time PCR technology. Cells lacking both Bright and Btk were transfected with Bright and/or Btk along with an immunoglobulin heavy chain reporter construct. Immunoglobulin gene transcription was enhanced when Bright and Btk were coexpressed. In contrast, neither Bright nor Btk alone led to activation of heavy chain transcription. Furthermore, Bright function required both Btk kinase activity and sequences within the pleckstrin homology domain of Btk. Bright was not appreciably phosphorylated by Btk; however, a third tyrosine-phosphorylated protein coprecipitated with Bright. Thus, the ability of Bright to enhance immunoglobulin transcription critically requires functional Btk.

ACKNOWLEDGMENTS

We thank Amy Windell for technical assistance and P. W. Tucker (Austin, Tex.), M. Coggeshall, and L. Thompson (OMRF) for critically reading the manuscript. We would also like to thank Kerry Humphrey for assistance preparing the manuscript.

This work was supported by National Institutes of Health grants AI044215 and AI45864 (C.F.W.) and T32 AI07633 (J.C.N.).