During this fiscal year we continued our studies to characterize the mechanisms that recruit and restrict the activation induced cytidine deaminase (AID) and its accompanying error-prone DNA repair machinery specifically to the immunoglobulin (Ig) genes. Our model system remains the DT40 cell line, a chicken B-cell line constantly undergoing somatic hypermutation (SHM) and Ig gene conversion (GCV), and showing the unique feature of being modifiable by standard gene targeting strategies. We were able to show mutational enhancer elements (MEEs) mediate the targeting of of SHM/GCV to Ig gene loci. Importantly, these MEEs are distinct from transcriptional enhancer elements. Furthermore we identified two such MEEs in the chicken IGL locus, and were able to narrow down one of them to a small DNA fragment about 200 bp in size. This element is evolutionarily conserved in terms of sequence and function. Lastly, we were able to show that the murine Ig kappa enhancers, that were long thought to act as targeting elements for SHM, solely serve as transcriptional enhancers. This in turns suggests that the targeting elements for SHM reside in as of yet uncharacterized non-coding DNA within the murine Ig kappa gene locus. Overall, our studies will provide a framework to explain the multiple levels at which the targeted introduction of mutations into Ig genes is controlled to protect the rest of the genome from potentially deleterious and cancer promoting alterations.
| Sen, Ranjan; Fugmann, Sebastian D (2012) Transcription, splicing, and release: are we there yet? Cell 150:241-3 |
| Kothapalli, Naga Rama; Fugmann, Sebastian D (2011) Targeting of AID-mediated sequence diversification to immunoglobulin genes. Curr Opin Immunol 23:184-9 |
| Kothapalli, Naga Rama; Norton, Darrell D; Fugmann, Sebastian D (2011) Classical Mus musculus Igýý enhancers support transcription but not high level somatic hypermutation from a V-lambda promoter in chicken DT40 cells. PLoS One 6:e18955 |
| Kothapalli, Nagarama; Fugmann, Sebastian D (2011) Characterizing somatic hypermutation and gene conversion in the chicken DT40 cell system. Methods Mol Biol 748:255-71 |
| Kothapalli, Naga Rama; Collura, Kaitlin M; Norton, Darrell D et al. (2011) Separation of mutational and transcriptional enhancers in Ig genes. J Immunol 187:3247-55 |
| Delker, Rebecca K; Fugmann, Sebastian D; Papavasiliou, F Nina (2009) A coming-of-age story: activation-induced cytidine deaminase turns 10. Nat Immunol 10:1147-53 |
