AID Binds Cooperatively with UNG and Msh2-Msh6 to Ig Switch Regions Dependent upon the AID C Terminus

Abstract

Activation-induced cytidine deaminase (AID) is induced in B cells during an immune response and is essential for both class switch recombination (CSR) and somatic hypermutation (SHM) of antibody genes. The C terminal 10 amino acids of AID are required for CSR but not for SHM, although their role in CSR is unknown. Using retroviral transduction into mouse splenic B cells, we show that the C terminus is not required for S region DSBs, and therefore functions downstream of DSBs. Using chromatin immunoprecipitation, we show that AID binds cooperatively with UNG and the mismatch repair proteins Msh2-Msh6 to Ig Sμ and Sγ3 regions, and this depends on the C terminus and the deaminase activity of AID. We also show that mismatch repair does not contribute to the efficiency of CSR in the absence of the AID C terminus. Although it has been demonstrated that both UNG and Msh2-Msh6 are important for introduction of S region DSBs, our data suggest that the ability of AID to recruit these proteins is important for DSB resolution, perhaps by directing the S region DSBs toward accurate and efficient CSR via non-homologous end joining.

Differential regulation of S-region hypermutation and class-switch recombination by noncanonical functions of uracil DNA glycosylase

“…2A), suggesting that increased UNG binding occurs upon AID-induced damage response in agreement with previous reports (32,36). Next, we examined the effect of UNG deficiency on the recruitment of Polβ, XRCC1, FEN1, and PARP1, after confirming their normal expression in UNG −/− cells (Fig.…”

A Structural Basis for the Biochemical Behavior of Activation-induced Deoxycytidine Deaminase Class-switch Recombination-defective Hyper-IgM-2 Mutants

“…Biochemical Properties of AID C-terminal Deletion MutantsThe C-terminal region binds proteins targeting AID to Ig switch region DNA (28,29). Two naturally occurring mutations causing a loss of 9 and 17 C-terminal amino acids of AID (⌬C 9 and ⌬C 17 ) were found in HIGM-2 patients (14); the patient with ⌬C 17 also has an insertion in the C-terminal region (14).…”

“…It has been suggested that the C terminus is essential for binding proteins such as UNG, MSH2, 14-3-3, and DNA-PK cs required to target AID to switch-region (28,29,36). For the HIGM-2 C-terminal mutants, although CSR is absent, SHM is almost normal (14,30), suggesting that AID retains a considerable level of C deamination activity when as many as 17 C-terminal amino acids have been removed.…”

“…It is well established that the C-terminal region is needed for binding proteins that target AID to Ig switch regions DNA (28,29,36). Deletions in this region cause a loss of CSR, but not SHM (14,30).…”

Separation of Function between Isotype Switching and Affinity Maturation In Vivo during Acute Immune Responses and Circulating Autoantibodies in UNG-Deficient Mice

“…Our results in vitro and in vivo indicate that UNG is not rate limiting for CSR. Interestingly, published experiments using the same retroviral delivery strategy and assay as us showed that overexpressed AID in Aid 2/2 Msh2 2/2 B cells produced IgG1 CSR levels that were ∼50% of those it produced in Aid 2/2 B cells (39), raising the possibility that MSH2/MSH6 could be rate limiting.…”