Identification and characterization of a positive regulatory region for activation induced cytidine deaminase mediated gene conversion in chicken B cells

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Title: Identification and characterization of a positive regulatory region for activation induced cytidine deaminase mediated gene conversion in chicken B cells
Author: Kim, Yonghwan, 1975-
Abstract: B cells have unique machinery to make up a large pool of antibody repertoire . After V (D )J recombination in early B cell development , the rearranged immunoglobulin genes are further diversified by somatic hypermutation (SHM ) , gene conversion (GC ) and class switch recombination (CSR ) . Acitvation induced cytidine deaminase (AID ) is a key initiating factor for SHM , GC and CSR . A majority of research data supports the model that AID modifies Ig genes at the DNA level by deaminating cytosines to uracils . The mutagenic activity of AID is largely restricted to Ig genes to avoid genomic instability in general . The specificity cannot be attributed to the primary sequence of the Ig genes since unrelated DNA is mutated by AID in the context of Ig genes . A clue to this problem is that AID function is dependent on transcription . Since not all transcribed genes are mutated by AID , there must be something special about the transcription of Ig genes , and the reasoning has prompted extensive analysis of Ig promoters and enhancers . We addressed this question in chicken B cell line DT40 . We identified a 2 .4 -kilobase regulatory region which is important for AID function both within and outside of Ig locus . This regulatory region contains binding sites for multiple transcription factors . Mutation of these binding sites impairs AID mediated gene conversion . In addition , ablation of NF -κB family member , c -Rel and p50 , reduces the AID targeting function of this regulatory region . Since the implicated transcription factors have been reported to associate with histone acetylases , the regulatory region may function by facilitating the access of AID to target DNA . To test this hypothesis , we used the I -SceI endonuclease and dam methylase as probes for chromatin structure . We found that the regulatory region does not increase chromatin accessibility to these probes . In fact , the regulatory region appears to interfere with the cleavage of target DNA by I -SceI . Another possible role of the regulatory region could be direct recruitment of AID to Ig genes . To test this hypothesis , we utilized Dam identification method . Surprisingly , we found that the regulatory region facilitates AID targeting to the Igλ locus .
URI: http : / /hdl .handle .net /2152 /ETD -UT -2009 -12 -674
Date: 2010-08-23

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Identification and characterization of a positive regulatory region for activation induced cytidine deaminase mediated gene conversion in chicken B cells. Doctoral dissertation, The University of Texas at Austin. Available electronically from http : / /hdl .handle .net /2152 /ETD -UT -2009 -12 -674 .

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