ChIP-seq of plasma cell-free nucleosomes identifies cell-of-origin gene expression programs

Abstract

Blood cell-free DNA (cfDNA) is derived from fragmented chromatin in dying cells. As such, it remains associated with histones that may retain the covalent modifications present in the cell of origin. Until now this rich epigenetic information carried by cell-free nucleosomes has not been explored at the genome level. Here, we perform ChIP-seq of cell free nucleosomes (cfChIP-seq) directly from human blood plasma to sequence DNA fragments from nucleosomes carrying specific chromatin marks. We assay a cohort of healthy subjects and patients and use cfChIP-seq to generate rich sequencing libraries from low volumes of blood. We find that cfChIP-seq of chromatin marks associated with active transcription recapitulates ChIP-seq profiles of the same marks in the tissue of origin, and reflects gene activity in these cells of origin. We demonstrate that cfChIP-seq detects changes in expression programs in patients with 1 heart and liver injury or cancer. cfChIP-seq opens a new window into normal and pathologic tissue dynamics with far-reaching implications for biology and medicine. One Sentence Summary:Chromatin immunoprecipitation and sequencing of histone modifications on blood-circulating nucleosomes (cfChIP-seq) provides detailed information about gene expression programs in different human organs.

Phenotypes from cell-free DNA

“…Protein-bound DNA lasts longer than naked DNA in serum [19], where nucleases are abundant, which suggests that cfDNA is probably double-stranded and protein-bound to inhibit degradation by endogenous nucleases in plasma. This is supported by the ability to detect nucleosomes in plasma by sandwich enzyme-linked immunosorbent assays (ELISA) and by chromatin immunoprecipitation (ChIP) [20][21][22][23][24]. Apoptosis is one of the processes that could lead to genome fragmentation into small protein-DNA complexes that are found in circulation.…”

“…Often promoter hypermethylation of a gene is associated with silencing [79]. DNA methylation profiles are stable and cell-type specific, plasma-derived cfDNA types TF-bound <50 bp TF occupancy [11,56] TSS analysis + copy-number gain [34] sub-nucleosome enrichment [10] window protection score/fast Fourier transformation [11] ELISA ChIP cfChIP-seq [19,[20][21][22][23] royalsocietypublishing.org/journal/rsob Open Biol. 10: 200119…”

“…A recent study sought to assess circulating post-translationally modified nucleosomes in a variety of physiological states using a combination of ChIP and next-generation sequencing [ 21 ]. The authors created a workflow to isolate modified circulating nucleosomes from plasma and performed high-throughput sequencing of the associated DNA (cfChIP-seq).…”

EGFR transcription in non‐small‐cell lung cancer tumours can be revealed in ctDNA by cell‐free chromatin immunoprecipitation (cfChIP)

“…KRT6 analysis [16]. Moreover, Sadeh et al [20] presented the cfChIP-seq method, which identifies transcriptionally active genes by next-generation sequencing. In addition, the study demonstrates cfChIPseq can identify the transcriptional profile of the various tissues contributing to the cfDNA pool [20].…”

Liquid Biopsies to Evaluate Immunogenicity of Gynecological/Breast Tumors: On the Way to Blood-Based Biomarkers for Immunotherapies

“…Most studies elucidating TMB as ICI biomarker use tumor tissue biopsies, but cfDNA and probably also CTCs, can be suitable LB sources for serial TMB assessment. Employing chromatin immunoprecipitation sequencing, the variants of tumor-derived cfDNA (ctDNA) and variants of cfDNA, released by non-pathogenic cells, might even be differentiated [78].…”