Dynamic Regulation of TET2 by Phosphorylation and Dephosphorylation at Ser99

Abstract

The ten‐eleven translocation enzyme was discovered when an 8‐year‐old boy diagnosed with acute myeloid leukemia (AML) containing the t(10;11)(q22;q23) in 2003. Three members, TET1, TET2 and TET3, belong to this dioxygenase family of enzymes. TET2 is widely implicated in cancer and in aging due to its role as a global epigenetic modifier catalyzing oxidation of 5‐methylcytosine (5mC) to 5‐hydroxymethylcytosine (5hmC), 5‐formylcytosine (5fC) and then 5‐carboxylcytosine (5caC) in 2‐oxogluterate (2‐OG) dependent manner. TET2 mutations are prevalent in malignancies including acute AML. Notably, most TET2 mutations in AML are monoallelic. Given the presence of a remaining wild type allele in these malignancies, characterizing the mechanism that regulate TET2 protein could have therapeutic significance.Human TET2 is 2002 amino acids in length. The catalytic domain is located in the C‐terminal portion of the protein between amino acids 1129–1936. The C‐terminal catalytic domain comprises a cysteine rich domain (1129–1312 amino acids) followed by a double stranded β‐helix domain (DSBH) which make a compact fold to execute the catalytic activity. The C‐terminal part of TET2 has been reported capable of oxidizing 5‐mC to 5‐hmC independent of the N‐terminus. As such, the function of the N‐terminus of this large protein remains poorly characterized.Here, we identify that several isoforms of the 14‐3‐3 family of proteins bind TET2. 14‐3‐3s bind TET2 when phosphorylated at serine 99 (S99). AMPK‐mediated phosphorylation at S99 promotes TET2 stability and increases global DNA 5‐hydroxymethylcytosine. 14‐3‐3s’ interaction with TET2 serves to protect S99 phosphorylation. Disruption of this interaction leads to both reduced TET2 phosphorylation and decreased protein stability. Furthermore, we identify that the protein phosphatase 2A (PP2A) can interact with TET2 and dephosphorylates S99. siRNA knockdown of PP2A (B‐alpha subunit) or pharmacologic inhibition of PP2A was able to significantly increase TET2 phosphorylation and protein level as well. Collectively, our study provides novel insights into the role of the N‐terminal domain in TET2 regulation. Moreover, they demonstrate the dynamic nature of TET2 protein regulation that could have therapeutic implications for disease states resulting from reduced TET2 levels and/or activity.Support or Funding InformationThe research reported in this article was supported by R01CA200653, I01BX002930, and a Mike Slive Foundation Grant (to S.S.) and in part by the UAB Comprehensive Cancer Center (P30CA013148).