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

Abstract

Activation-induced deaminase converts deoxycytidine to deoxyuridine at the Ig loci. Complementary pathways, initiated by the uracil-DNA glycosylase (UNG) or the mismatch repair factor MSH2/MSH6, must process the deoxyuridine to initiate class-switch recombination (CSR) and somatic hypermutation. UNG deficiency most severely reduces CSR efficiency and only modestly affects the somatic hypermutation spectrum in vitro. This would predict isotype-switching deficiency but normal affinity maturation in Ung−/− mice in vivo, but this has not been tested. Moreover, puzzling differences in the amount of circulating Ig between UNG-deficient humans and mice make it unclear to what extent MSH2/MSH6 can complement for UNG in vivo. We find that Ab affinity maturation is indeed unaffected in Ung−/− mice, even allowing IgM responses with higher than normal affinity. Ung−/− mice display normal to only moderately reduced basal levels of most circulating Ig subclasses and gut-associated IgA, which are elicited in response to chronically available environmental Ag. In contrast, their ability to produce switched Ig in response to immunization or vesicular stomatitis virus infection is strongly impaired. Our results uncover a specific need for UNG in CSR for timely and efficient acute Ab responses in vivo. Furthermore, Ung−/− mice provide a novel model for separating isotype switching and affinity maturation during acute (but not chronic) Ab responses, which could be useful for dissecting their relative contribution to some infections. Interestingly, Ung−/− mice present with circulating autoantibodies, suggesting that UNG may impinge on tolerance.

APE1 is dispensable for S-region cleavage but required for its repair in class switch recombination

“…More importantly, APE1's endonuclease activity is not required for the AIDinduced DNA cleavage, in parallel with the findings that UNG is dispensable for the DNA cleavage step during SHM (56,57) and CSR (39,58). Recently, we reported that UNG enhances AIDdependent S-S synapse formation by recruiting p53-binding protein 1 and DNA-dependent protein kinase, catalytic subunit (49).…”

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

“…We also confirmed that a UNG mutant with the N-terminal 90 residues deletion (Δ90 UNG) showed similar or even better CSR rescue, albeit slightly less s-SHM suppression, indicating that the core domain of UNG plays a major role in mutation suppression as well as CSR promotion. Consistent with our finding, recent observation by Zahn et al (33) revealed that UNG-deficient mice have even stronger SHM in the V region than WT. The N terminus of UNG is unique in that it possesses proliferating cell nuclear antigen (PCNA) and replication protein A (RPA) interacting sites (see Fig.…”

“…If UNG is involved in AID-dependent DNA cleavage, UNG deficiency is supposed to reduce not only CSR but also SHM because SHM is not restricted C/G and depends on error-prone repair of DNA breakage (12). Surprisingly, however, careful data analyses indicate that UNG deficiency rather augments SHM, not only in Ig loci but also in other target loci such as c-myc and bcl-6 (30,(32)(33)(34).…”

Activation induced deaminase C-terminal domain links DNA breaks to end protection and repair during class switch recombination

“…However, despite the importance of UNG in generating DSBs for CSR (3), breaks have been found to happen with similar frequency at the Sμ of murine B cells expressing ΔE5 and R190X compared with AID (13,14) as well as in AICDA R190X/+ vs. AICDA +/+ human B cells (15). Because we see ∼50% reduction in Sμ occupancy by UNG after ΔE5, it may be that UNG is not limiting, which was suggested by the normal CSR Ung haploinsufficient mice and B cells (43). Alternatively, mismatch repair enzymes could compensate for UNG (26), although whether ΔE5 recruits more or less MSH2 to the Sμ is unclear (13,42).…”