Cell cycle-specific UNG2 phosphorylations regulate protein turnover, activity and association with RPA

Abstract

Human UNG2 is a multifunctional glycosylase that removes uracil near replication forks and in non-replicating DNA, and is important for affinity maturation of antibodies in B cells. How these diverse functions are regulated remains obscure. Here, we report three new phosphoforms of the non-catalytic domain that confer distinct functional properties to UNG2. These are apparently generated by cyclin-dependent kinases through stepwise phosphorylation of S23, T60 and S64 in the cell cycle. Phosphorylation of S23 in late G1/early S confers increased association with replication protein A (RPA) and replicating chromatin and markedly increases the catalytic turnover of UNG2. Conversely, progressive phosphorylation of T60 and S64 throughout S phase mediates reduced binding to RPA and flag UNG2 for breakdown in G2 by forming a cyclin E/c-myc-like phosphodegron. The enhanced catalytic turnover of UNG2 p-S23 likely optimises the protein to excise uracil along with rapidly moving replication forks. Our findings may aid further studies of how UNG2 initiates mutagenic rather than repair processing of activation-induced deaminase-generated uracil at Ig loci in B cells.

Cyclin-Dependent Kinases Regulate Ig Class Switching by Controlling Access of AID to the Switch Region

“…Additionally, CDKs may also be involved in the steps following AID initiating the CSR. Previous studies have suggested that UNG activity is regulated by CDK-dependent phosphorylation (35). Therefore, inhibition of CDK activity may compromise UNG activity and in turn result in reduced class switching.…”

Uracil in DNA and its processing by different DNA glycosylases

“…Furthermore, the triple phosphorylated form is mono-ubiquitinylated late in the S-phase and then rapidly degraded in the G2-phase. Interestingly, the sequence encompassing the phosphorylated Thr60-Ser64 forms a motif that is very similar to a phosphodegron, probably explaining the rapid degradation in G2 (Hagen et al 2008). Why UNG2 must be degraded remains obscure, but some observations indicate a toxicity of unregulated expression of the enzyme (Henrik Sahlin Pettersen 2007, unpublished data).…”

Competitive repair pathways in immunoglobulin gene hypermutation

“…AID expression is indeed not cell-cycle regulated, making it likely that cytosine deamination may take place at different phases of the cycle . By contrast, UNG is strongly upregulated in the late G1 and early S-phases, at which stage it can cope with misincorporated uracils during replication (Hagen et al 2008). MMR would be normally considered as a post-replicative actor, but the restricted involvement of the sole MSH2-MSH6 and Exo1 partners precludes such a priori.…”