The old saying “every pot will find its lid” suggests that a match exists for everyone looking for love, you just have to get out there and find your perfect partner! Online-dating may help scientists to find their significant other from the bench side; however, current laboratory approaches for the identification of protein-binding partners for specific epigenetic modifications in living cells can leave any researcher a little broken-hearted.
Now, a romantic team of researchers led by Tuncay Baubec (University of Zurich, Switzerland) sought to act as master matchmakers between “epigenetic pots” and “protein lids” in a recent study that described the development of “ChromID” – a method to identify the local protein composition at regions marked by individual and combinatorial epigenetic marks.
So, let’s allow Villaseñor and colleagues to tell their starry-eyed story of romance that employs mouse embryonic stem cells as a positively dreamy model system to match epigenetic pots to protein lids:
- The authors first created biotin- (purification)
and GFP- (intracellular tracking) labeled engineered chromatin reader (eCR)
proteins specific for four different epigenetic modifications
- The identification of H3K9me3, DNA methylation (5-methylcytosine), H3K4me3, and H3K27me3 employs the chromodomain from CBX1, the MBD domain from MBD1/MeCP2, the chromodomain from CBX7, and the Phd domain from TAF3, respectively, as the integral part of each eCR
- The high specificity of these four single-modification eCRs, as characterized and validated by chromatin immunoprecipitation and mass spectrometry, highlights their suitability as multipurpose probes for the detection of chromatin modification patterns in living cells
- The eCRs are then fused to a
promiscuous biotin ligase (BASU) to create ChromID as a novel means to
identify proteins that bind to specific epigenetic modifications based on
- ChromID for each epigenetic modification permits the identification of expected interacting proteins and also a raft of novel binding proteins
- As an example, H3K9me3 ChromID identified both high-confidence proteins, such as H3K9 methyltransferases, heterochromatin-associated proteins, chromatin remodelers, and novel binding proteins, including CASZ1, ZNF24, ZNF292, ZNF512B, ZNF518B, ZNF280B, and ZNF280D
- Finally, the authors constructed a combinatorial
ChromID system composed of eCRs for both H3K4me3 and H3K27me3 to understand the
protein composition of the pluripotency-associated bivalent
- Bivalent ChromID identified 33 high-confidence factors associated with bivalent chromatin, including basal transcription factors, histone acetyltransferases, polycomb proteins, histone lysine demethylases, PRC1/2 components, and methylcytosine dioxygenases
The authors of this adoring article highlight how the use of natural reader domains to study the binding of proteins to specific modifications in live cells does away with problems such as the requirement for crosslinking/hybridization and the use of antibodies, whose variable affinity and avidity, lack of availability, and high cost that have often hindered research in this area. Furthermore, looking longingly to the future, this team of matchmakers also underscores the potential of ChromID to chart the epiproteome during dynamic cellular processes and development via tissue-specific experimentation in living animals.
Don’t despair; your perfect match may be just a click away! Find the perfect protein lid for your epigenetic pot over at Nature Biotechnology, March 2020.