Repetitive DNA sequences in the immunoglobulin switch mu region form RNA-containing secondary structures and undergo hypermutation by activation-induced deaminase (AID). To examine how DNA structure affects transcription and hypermutation, we mapped the position of RNA polymerase II molecules and mutations across a 5 kb region spanning the intronic enhancer to the constant mu gene. For RNA polymerase II, the distribution was determined by nuclear run-on and chromatin immunoprecipitation assays in B cells stimulated ex vivo. RNA polymerases were found at a high density in DNA flanking both sides of a 1 kb repetitive sequence that forms the core of the switch region. We propose that altered DNA structure in the switch region pauses RNA polymerase II. We are currently examining secondary structure and transcription patterns in the variable region, using knock-in mice containing one rearranged variable gene on both alleles. RNA polymerase II pausing is measured by nuclear run-on and ChIP techniques.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIAAG000714-06
Application #
8736595
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
6
Fiscal Year
2013
Total Cost
$609,058
Indirect Cost
Name
National Institute on Aging
Department
Type
DUNS #
City
State
Country
Zip Code
Rajagopal, Deepa; Maul, Robert W; Ghosh, Amalendu et al. (2009) Immunoglobulin switch mu sequence causes RNA polymerase II accumulation and reduces dA hypermutation. J Exp Med 206:1237-44
Gearhart, Patricia J; Sen, Ranjan (2008) Regulating antibody diversity: taming a mutagen. Mol Cell 31:615-6