title: The AID antibody diversification enzyme is regulated by protein kinase A phosphorylation.
abstract: Antibodies, which are produced by B-lineage cells, consist of immunoglobulin heavy (IgH) and light (IgL) chains that have amino-terminal variable regions and carboxy-terminal constant regions. In response to antigens, B cells undergo two types of genomic alterations to increase antibody diversity. Affinity for antigen can be increased by introduction of point mutations into IgH and IgL variable regions by somatic hypermutation. In addition, antibody effector functions can be altered by changing the expressed IgH constant region exons through IgH class switch recombination (CSR). Somatic hypermutation and CSR both require the B-cell-specific activation-induced cytidine deaminase protein (AID), which initiates these reactions through its single-stranded (ss)DNA-specific cytidine deaminase activity. In biochemical assays, replication protein A (RPA), a ssDNA-binding protein, associates with phosphorylated AID from activated B cells and enhances AID activity on transcribed double-stranded (ds)DNA containing somatic hypermutation or CSR target sequences. This AID-RPA association, which requires phosphorylation, may provide a mechanism for allowing AID to access dsDNA targets in activated B cells. Here we show that AID from B cells is phosphorylated on a consensus protein kinase A (PKA) site and that PKA is the physiological AID kinase. Thus, AID from non-lymphoid cells can be functionally phosphorylated by recombinant PKA to allow interaction with RPA and promote deamination of transcribed dsDNA substrates. Moreover, mutation of the major PKA phosphorylation site of AID preserves ssDNA deamination activity, but markedly reduces RPA-dependent dsDNA deamination activity and severely impairs the ability of AID to effect CSR in vivo. We conclude that PKA has a critical role in post-translational regulation of AID activity in B cells.
authors.affiliation: The Howard Hughes Medical Institute, The Children's Hospital, The CBR Institute for Biomedical Research, and Department of Genetics, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: Uttiya
authors.initials: U
authors.lastname: Basu
authors.forename: Jayanta
authors.initials: J
authors.lastname: Chaudhuri
authors.forename: Craig
authors.initials: C
authors.lastname: Alpert
authors.forename: Shilpee
authors.initials: S
authors.lastname: Dutt
authors.forename: Sheila
authors.initials: S
authors.lastname: Ranganath
authors.forename: Gang
authors.initials: G
authors.lastname: Li
authors.forename: Jason Patrick
authors.initials: JP
authors.lastname: Schrum
authors.forename: John P
authors.initials: JP
authors.lastname: Manis
authors.forename: Frederick W
authors.initials: FW
authors.lastname: Alt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000201:enzymology / Q000378:metabolism; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D003564:Cytidine Deaminase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D011485:Protein Binding; D051716:Replication Protein A / Q000378:metabolism; D027041:Somatic Hypermutation, Immunoglobulin; D000098830:AICDA (Activation-Induced Cytidine Deaminase)
chemical_list: D051716:Replication Protein A; D017868:Cyclic AMP-Dependent Protein Kinases; D000098830:AICDA (Activation-Induced Cytidine Deaminase); D003564:Cytidine Deaminase