Targeted bisulfite sequencing of analog epigenetic memory in mammalian cells

Cells store information in the chromatin state by means of chemical modifications to DNA and histones that self-propagate through cell divisions and time. Although these modifications can keep gene expression silenced over generations, whether and how they maintain any other gene expression state remains unknown. Here, we found that chromatin modifications can maintain a wide range of gene expression levels over time, a phenomenon that we call analog epigenetic memory. We engineered a genomic reporter and epigenetic effectors to monitor with clonal resolution the gene expression dynamics following targeted perturbations to the chromatin state. Any gene expression level resulting from these perturbations stably persisted over time along with a distinct grade of DNA methylation. Altering the DNA methylation grade directly or interfering with DNA methylation maintenance resulted in permanent loss of gene expression memory. This identifies DNA methylation as the chief chromatin mark enabling analog epigenetic memory of gene expression. Consistent with the experiments, our chromatin modification model indicates that analog memory requires that the positive feedback between DNA methylation and Histone H3 Lysine 9 trimethylation (H3K9me3) is sufficiently weak. Our discovery will lead to a deeper understanding of epigenetic memory and to new tools for synthetic biology.