High-throughput single-cell DNA methylation and chromatin accessibility co-profiling with SpliCOOL-seq

Area of Science:

• Epigenetics
• Genomics
• Cancer Biology

Background:

• DNA methylation and chromatin accessibility are key epigenetic regulators influencing gene expression and cellular identity.
• Dysregulation of these epigenetic marks is implicated in cancer development and progression.
• Existing single-cell multi-omics technologies face limitations in throughput and sensitivity, restricting comprehensive biomarker discovery.

Purpose of the Study:

• To develop a high-throughput single-cell sequencing technology for simultaneous profiling of DNA methylation and chromatin accessibility.
• To enhance sensitivity and scalability for multi-modal epigenetic analysis in thousands of cells.
• To enable novel biomarker discovery and deepen the understanding of cancer epigenetics.

Main Methods:

• Development of single-cell split-pool ligation-based multi-omics sequencing technology (SpliCOOL-seq).
• Integration of in situ GpC methylation, universal Tn5 tagmentation, and split-pool combinatorial barcoding.
• High-throughput profiling of whole-genome DNA methylation and chromatin accessibility in single cells.

Main Results:

• SpliCOOL-seq accurately differentiated lung cancer cell types using genetic and epigenetic data.
• DNA methyltransferase (DNMT) inhibitors (5-Azacitidine, Decitabine) induced distinct large-scale demethylation patterns.
• Identification of novel DNA methylation biomarkers (e.g., FAM124B, SFN, OR7E47P) associated with patient survival in lung adenocarcinoma.
• Revealed accelerated epigenetic ageing and mitotic activity in tumor subclones, offering insights into tumorigenesis.

Conclusions:

• SpliCOOL-seq enables high-throughput, parallel profiling of DNA methylation and chromatin accessibility at single-cell resolution.
• The technology serves as a powerful platform for discovering cancer biomarkers and therapeutic targets.
• Applications revealed novel diagnostic markers and potential therapeutic targets, advancing precision oncology.