Changes in immunoglobulin-nucleoprotein complex structure mapped by chromatin immunoprecipitation

Daniel C. McDevit, Barbara S. Nikolajczyk

Research output: Contribution to journalArticlepeer-review

1 Scopus citations


Transcription factor-mediated immunoglobulin (Ig) enhancer activation has been analyzed extensively outside the physiological constraints of chromatin. Towards understanding the role sequence-specific DNA binding proteins identified by these methods play in activating Ig genes during B cell development, we have investigated in vivo interaction between the Ig enhancer activator PU.1 and two target elements, the Igμ and κ3′ enhancers, by chromatin immunoprecipitation (ChIP). By using two antibodies recognizing different PU.1 epitopes in murine B cells, these analyses demonstrate that ChIP results may depend on the availability of the epitope(s) targeted by the immunoprecipitating antibody. Specifically, PU.1 epitope availability at the μ and κ3′ enhancers does not accurately quantitate total PU.1 association. This result suggests the nucleoprotein complexes formed at these various active enhancers is cell type-specific. Interestingly, RAG1-/- but not RAG2-/- pro-B cells lack PU.1/κ3′ association, probably due to limited accessibility of the κ locus in the former. The more robust association of PU.1 with the κ3′ versus μ enhancer in all but RAG1-/- B lineage cells is not explained by differences in PCR primer efficiency, but likely reflects the different structures formed by the complexes at μ versus κ3′ enhancers. Finally, PU.1 is not associated with an inaccessible μ or κ3′ enhancer chromatin structure in macrophages, again emphasizing the importance cellular protein context plays in PU.1/Ig enhancer association. The demonstration that changes in epitope availability, hence nucleoprotein structure, can be monitored by ChIP suggests using this technique to monitor biologically important changes in nucleoprotein complex structure/composition in situ.

Original languageEnglish
Pages (from-to)1541-1548
Number of pages8
JournalMolecular Immunology
Issue number10
StatePublished - Apr 2006


  • B cell
  • Gene regulation
  • Molecular biology
  • Transcription factors

ASJC Scopus subject areas

  • Immunology
  • Molecular Biology


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