μDamID: a microfluidic approach for imaging and sequencing protein-DNA interactions in single cells

Abstract

1 2 Genome regulation depends on carefully programmed protein-DNA interactions that maintain or 3 alter gene expression states, often by influencing chromatin organization. Most studies of these 4 interactions to date have relied on bulk methods, which in many systems cannot capture the 5 dynamic single-cell nature of these interactions as they modulate cell states. One method allowing 6 for sensitive single-cell mapping of protein-DNA interactions is DNA adenine methyltransferase 7 identification (DamID), which records a protein's DNA-binding history by methylating adenine 8 bases in its vicinity, then selectively amplifies and sequences these methylated regions. These 9interaction sites can also be visualized using fluorescent proteins that bind to methyladenines. Here 10 we combine these imaging and sequencing technologies in an integrated microfluidic platform 11 (µDamID) that enables single-cell isolation, imaging, and sorting, followed by DamID. We apply 12 this system to generate paired single-cell imaging and sequencing data from a human cell line, in 13 which we map and validate interactions between DNA and nuclear lamina proteins, providing a 14 measure of 3D chromatin organization and broad gene regulation patterns. µDamID provides the 15 unique ability to compare paired imaging and sequencing data for each cell and between cells, 16 enabling the joint analysis of the nuclear localization, sequence identity, and variability of protein-17

μCB-seq: microfluidic cell barcoding and sequencing for high-resolution imaging and sequencing of single cells

“…65 For example, μCB-seq could be paired with super-resolution microscopy to investigate phase separation of superenhancers and its effect on gene expression across the whole transcriptome of individual cells. 66 Another implementation could pair μCB-seq with microfluidic DamID 67 to investigate the bidirectional interplay between gene expression and chromatin organization in the same single cell. We believe the μCB-seq platform will be a powerful tool for investigations aiming to understand the association between a phenotype and the transcriptome, thereby gaining a highresolution fingerprint for a particular cell population identified using other higher-throughput scRNA-seq protocols.…”