Replication protein A interacts with AID to promote deamination of somatic hypermutation targets.
Activation-induced cytidine deaminase (AID) is a single-stranded (ss) DNA deaminase required for somatic hypermutation (SHM) and class switch recombination of immunoglobulin genes. Class switch recombination involves transcription through switch regions, which generates ssDNA within R loops. However, although transcription through immunoglobulin variable region exons is required for SHM, it does not generate stable ssDNA, which leaves the mechanism of AID targeting unresolved. Here we characterize the mechanism of AID targeting to in-vitro-transcribed substrates harbouring SHM motifs. We show that the targeting activity of AID is due to replication protein A (RPA), a ssDNA-binding protein involved in replication, recombination and repair. The 32-kDa subunit of RPA interacts specifically with AID from activated B cells in a manner that seems to be dependent on post-translational AID modification. Thus, our study implicates RPA as a novel factor involved in immunoglobulin diversification. We propose that B-cell-specific AID-RPA complexes preferentially bind to ssDNA of small transcription bubbles at SHM 'hotspots', leading to AID-mediated deamination and RPA-mediated recruitment of DNA repair proteins.
Chaudhuri
Jayanta
J
Howard Hughes Medical Institute, Children's Hospital, Center for Blood Research and Department of Genetics, Harvard University Medical School, Boston, Massachusetts 02115, USA.
Khuong
Chan
C
Alt
Frederick W
FW
D000586:Amination; D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D003564:Cytidine Deaminase / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D017353:Gene Deletion; D006801:Humans; D051379:Mice; D011485:Protein Binding; D051716:Replication Protein A; D027041:Somatic Hypermutation, Immunoglobulin; D013379:Substrate Specificity; D014158:Transcription, Genetic / Q000235:genetics; D000098830:AICDA (Activation-Induced Cytidine Deaminase)
D004268:DNA-Binding Proteins; C497167:RPA1 protein, human; D051716:Replication Protein A; C497168:Rpa1 protein, mouse; D004247:DNA; D000098830:AICDA (Activation-Induced Cytidine Deaminase); D003564:Cytidine Deaminase