Protein kinase A inhibits tumor mutator APOBEC3B through phosphorylation

Abstract

APOBEC3B cytidine deaminase (A3B) catalyzes cytosine into uracil in single-strand DNA and induces C-to-T mutations in genomic DNA of various types of tumors. Accumulation of APOBEC signature mutations is correlated with a worse prognosis for patients with breast cancer or multiple myeloma, suggesting that A3B activity might be a cause of the unfavorable DNA mutations and clonal evolution in these tumors. Phosphorylation of conserved threonine residues of other cytidine deaminases, activation induced deaminase (AID) and APOBEC3G, inhibits their activity. Here we show that protein kinase A (PKA) physically binds to A3B and phosphorylates Thr214. In vitro deaminase assays and foreign DNA editing assays in cells confirm that phosphomimetic A3B mutants, T214D and T214E, completely lose deaminase activity. Molecular dynamics simulation of A3B phosphorylation reveals that Thr214 phosphorylation disrupts binding between the phospho-A3B catalytic core and ssDNA. These mutants still inhibit retroviral infectivity at least partially, and also retain full anti-retrotransposition activity. These results imply that PKA-mediated phosphorylation inhibits A3B mutagenic activity without destructing its innate immune functions. Therefore, PKA activation could reduce further accumulation of mutations in A3B overexpressing tumors.

The effect of APOBEC3B deaminase on double-stranded DNA

“…Nonetheless, these results indicate that deaminase activity is dispensable for the effects mediated by A3B in MNNG-treated cells. Deaminase independent activity has been found in other instances of A3-mediated effects including A3B [40][41][42][43][44][45] . We also found that knockdown of endogenous A3B by siRNA did not affect the level of γ-H2AX foci ( Supplementary Fig.…”

“…A3B, like other members of the A3 family, is a single-strand specific DNA binding protein 44,55 . This property is presumably related to the numerous instances of A3 deaminase-independent effects [40][41][42][43][44][45] . Thus, our findings that A3B deaminase activity is not required for its aggravation of MNNGinduced DSBs suggest that it does so as a consequence of binding to the single-stranded regions that characterize MNNG-induced damage, perhaps by interfering with or delaying the eventual repair of these lesions.…”

Molecular basis of clonal evolution in multiple myeloma

“…Ras mutations are mostly subclonal and detected at the time of both diagnosis and relapse [16,18,22,36], consistent with their role in tumor progression but not the initiation of MM. Little is known about the mechanisms underlying the acquisition of non-synonymous point mutations in Ras family oncogenes by MM cells; however, the involvement of the APOBEC3B cytidine deaminase was suggested by a recent comprehensive analysis of the mutation signature of multiple cancers [37][38][39] and subsequent biochemical studies [40,41]. APOBEC3B confers hypermutator phenotypes to cancer cells by converting cytidine to uracil/thymidine in TCN trinucleotide repeats in the genome [40][41][42].…”

“…Little is known about the mechanisms underlying the acquisition of non-synonymous point mutations in Ras family oncogenes by MM cells; however, the involvement of the APOBEC3B cytidine deaminase was suggested by a recent comprehensive analysis of the mutation signature of multiple cancers [37][38][39] and subsequent biochemical studies [40,41]. APOBEC3B confers hypermutator phenotypes to cancer cells by converting cytidine to uracil/thymidine in TCN trinucleotide repeats in the genome [40][41][42]. This enzyme has been demonstrated to be a transcriptional target of Maf transcription factors and may contribute to rapid progression and treatment resistance of MM cases carrying t (14;16) and t (14;20) [37,42].…”

“…In this framework, high-risk abnormalities, such as deregulation of the Maf family transcription factors and the MMSET histone methyltransferase, may not be a simple consequence of disease progression but a driving force of the disease by accelerating clonal heterogeneity, ultimately leading to a clonal dominance of selected ones [14,15,62]. As mentioned above, transactivation of the APOBEC3B cytidine deaminase by Maf transcription factors provides a molecular basis for this concept [37][38][39][40][41]. The branching evolution is a Fig.…”