Elucidating IgH transcriptional regulation is critical for understanding how the humoral immune response is controlled. In addition, it provides an important paradigm for understanding tissue-specific and developmentally regulated gene transcription. The investigators past work has helped to identify and characterize proteins which regulate the transcriptional activity of VH promoters and the Eu enhancer. To continue studies on IgH transcriptional regulation, the investigator proposes experiments in three general areas: 1) to determine the biological importance of particular IgH transcription proteins in the context of B-cell development and function, 2) to identify the biochemical mechanisms of action of IgH transcription proteins and 3) to study the role of germline VH transcripts in VDJ recombination. Studies will focus primarily on C/EBP and uE3 binding proteins based on the investigator's past experience and because the play an important role in IgH transcription. To study the biological role of uE3 and C/EBP binding proteins, mice deficient in TFE3 or Ig/EBP will be created and studied. Long term bone marrow cultures will be exploited to study the roles of uE3 and C/EBP proteins in primary B-cell development and for lymphopoiesis in vivo. In vitro transcription utilizing reconstituted chromatin templates will be used to study the biochemical mechanism of action of uE3 and c/EBP proteins bound to both distal (enhancer) and proximal sites. RAG-/- cell lines will be studied to determine if germline VH transcripts are required for VDJ recombination and gene targeting will be employed to test whether transcription and/or altered chromatin structure are sufficient to target recombination of particular VH gene segments.
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