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Antibody diversification requires the DNA deaminase AID to induce DNA instability at immunoglobulin (Ig) loci upon B cell stimulation. For efficient cytosine deamination, AID requires single-stranded DNA and needs to gain access to Ig loci, with RNA pol II transcription possibly providing both aspects. To understand these mechanisms, we isolated and characterized endogenous AID-containing protein complexes from the chromatin of diversifying B cells. The majority of proteins associated with AID belonged to RNA polymerase II elongation and chromatin modification complexes. Besides the two core polymerase subunits, members of the PAF complex, SUPT5H, SUPT6H, and FACT complex associated with AID. We show that AID associates with RNA polymerase-associated factor 1 (PAF1) through its N-terminal domain, that depletion of PAF complex members inhibits AID-induced immune diversification, and that the PAF complex can serve as a binding platform for AID on chromatin. A model is emerging of how RNA polymerase II elongation and pausing induce and resolve AID lesions.

Original publication

DOI

10.1084/jem.20112145

Type

Journal article

Journal

J exp med

Publication Date

22/10/2012

Volume

209

Pages

2099 - 2111

Keywords

Animals, Antibody Diversity, B-Lymphocytes, Blotting, Western, Cell Line, Tumor, Chromatin, Cytidine Deaminase, DNA-Binding Proteins, HEK293 Cells, HeLa Cells, High Mobility Group Proteins, Humans, Immunoglobulin Class Switching, Immunoglobulins, Immunoprecipitation, Nuclear Proteins, Protein Binding, RNA Interference, Transcription Factors, Transcriptional Elongation Factors