Believers in the Sasquatch say deep mountain valleys are where they may be hiding. When it comes to our genomes, DNA methylation valleys hold secrets yet to be discovered too. How DNA methylation is coordinated at multiple CpGs sites across regulatory regions in early development is still poorly understood. Bivalent promoters are one system used to study this. These “poised” regions are marked with both active (H3K4me3) and repressive (H3K27me3) histone modifications. The presence of H3K4me3 means that DNA methylation should be mostly absent at these sites. Thus, the landscape of bivalent promoters often contains “DNA methylation valleys”: large regions of low DNA methylation.
The labs of Olivier Elemento and Danwei Huangfu wanted to discover regulators of DNA methylation at bivalent promoters in human embryonic stem cells (hESCs). To do this, they developed a genome-wide CRISPR Cas9 screen that utilized DNA methylation at the bivalent PAX6 gene as a reporter. They inserted the methylation sensitive Snrpn promoter to express the tdTomato fluorescent protein upstream of PAX6 in an hESC line. Thus, the fluorescence in the line represents the DNA methylation level at a bivalent promoter. They performed a genome-wide CRISPR-Cas9 screen using a lentivirial library to target all human genes for knockdown and observe the effect at PAX6 methyaltion. Here’s what they found:
- 54 gene (cell population) “sightings” that they selected for validation, which include genes known to affect DNA methylation: TET1, TDG, KDM2B, and BCOR
- QSER1 ranks highest in the screen; however, its function was unknown, so they focused on characterizing it
- QSER1 knockout causes hypermethylation similar to the degree and genomic location of TET1 knockout
- These changes mostly occur at regulatory regions (promoters and enhancers), especially bivalent promoters, PRC2 binding sites, and H3K27me3 peaks
- The genomic occupancy of QSER1 highly correlates with TET1, which led the team to hypothesize that they have mutual dependence, so they knocked out each and found that the binding of each was reduced in the absence of the other.
- QSER1 binding protects DNA methylation valleys from DNA methylation by preventing DNMT3A/B binding
- Combined knockout of QSER1 and TET1 has a larger effect on DNA methylation and gene expression than either alone, and prevents hESC differentiation.
Taken together, these data establish a role for QSER1 as an antagonist to DNA methylation at DNA methylation valleys and other important regions in early development. More broadly, it shows that there is still much to learn about epigenetic regulation, as unbiased screens for specific protein functions can still identify important, uncharacterized players. So, while the jury is still out on the Sasquatch, there are still unknown protein players lurking in valleys of our epigenome.
Get into all the peaks and valleys of this story in Science, April 2021