A romantic “tête-à-tête” between two lovebirds usually involves helping factors which set the mood and make the night move along slickly. A good wine, some soft lighting, and maybe even a little smooth jazz! While that might work for the humble researcher, the “TET-á-TET” involved in DNA demethylation is a slightly different affair and demands something a little more specific!
DNA demethylation itself involves a step-by-step oxidation process controlled by the Ten-eleven Translocation (TET) enzymes. Such enzymes, including TET1 and TET2, convert 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC), then into 5-formylcytosine (5fC), and finally into 5-carboxylcytosine (5caC). This process plays an integral role in the regulation of gene expression, amongst other processes, although which additional elements facilitate the where and the when of this DNA demethylation TET-á-TET remain relatively unknown.
This led researchers from the lab of Bing Zhu (Chinese Academy of Sciences, Beijing, China) to pick apart the process and determine the factors required for TET-mediated DNA demethylation in mouse embryonic stem cells (mESCs). In their new study, Xiong and Zhang et al now demonstrate that SALL4A is the factor that sets the mood for a DNA demethylation TET-á-TET!
Initial in vitro analysis combined quantitative proteomic technique (SILAC – Stable Isotope Labeling by/with Amino acids in Cell culture) with DNA affinity purification to discover what DNA binding proteins liked to bind 5hmC residues. This provided a list of known 5hmC binding factors, such as UHRF1 and MeCP2, but highlighted that the SALL4A zinc finger protein (highly expressed in mESCs) also had a very strong attraction for 5hmC residues.
Subsequent in vivo chromatin immunoprecipitation (ChIP) analysis in mESCs demonstrated that SALL4A and TET1 co-bound at enhancers, which are recognized “hot-spots” for oxidized 5mC residues such as 5hmC. Further analysis then discovered that SALL4A also permitted TET2 to enter the foray and convert 5hmC into their more oxidized forms (5fC and 5caC) to permit full DNA demethylation. SALL4A loss inhibited enhancer association of TET2 and stopped the step-by-step oxidation of 5hmC, so pointing to the important organizational properties of SALL4A.
Overall, the authors suggest that SALL4A sets the mood and makes the DNA demethylation TET-á-TET a process to remember! However, the study does note that yet more supporting factors are likely to hiding in the shadows awaiting discovery (champagne? caviar nibbles? OCT4? NANOG?), and furthermore, that other regulators related to SALL4A may exist and exert related DNA demethylation regulatory functions at other genomic locations and in other cell types.
But for now, sit back and relax and have a TET-á-TET or your own – you and a copy of this excellent new study! Read all about it at Mol Cell, Nov 2016.