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Scientists Develop Method of Detecting DNA Regulatory Sites in Single Cells: Study Reduces Need for Millions of Cells in Some Analyses

Scientists have discovered a method for genome-wide detection of DNA regulatory sites by using single cells instead of millions of cells, which have been needed in older techniques. DNA regulatory sites help control gene expression — that is, the different types of proteins a cell makes. Gene expression, in turn, controls much of the character and activities of the cell — both normal and pathobiological processes.

The new technique, described in a recent article in Nature, helps scientists target specific regions called DNase I hypersensitive sites (DHSs), key genomic regulatory elements where chromatin — the complexes of DNA and proteins constituting chromosomes — is no longer condensed. In DHSs, chromatin is more extended and the enzyme DNase I can be used to essentially break open, or cleave, the DNA molecule exposing its regulatory regions.  

The conventional method requiring millions of cells for mapping DHSs is called DNase sequencing (DNase-seq). The new technique, referred to as single-cell DNase sequencing (scDNase-seq), was developed by a research team led by Keji Zhao, PhD, of the Laboratory of Epigenome Biology at the National Heart, Lung, and Blood Institute (NHLBI).

“Our method uses a circular DNA molecule as a buffer to increase recovery of DHS DNA during the preparation of what are called ‘DNA libraries,’ which are like copies of the genomic DNA being studied,” said Dr. Zhao. “On average, we attained 317,000 unique scDNase-seq reads and 38,000 DHSs could be identified in each single-cell library.”

With the detailed information it can give for identifying regulatory regions per individual cell, scDNase-seq has the potential to provide greater clarity regarding cellular activities. It also has the potential of generating critical information for precision medicine, in which medical decisions, practices, and products are tailored to the individual patient to help optimize care.

“Scientists have been able to detect genome-wide DHSs using DNase-seq for several years,” said study investigator Wenfei Jin, PhD, of the Systems Biology Center at NHLBI. “However, the millions of cells needed for preparing a conventional DNase-seq library blocked the use of DHS analyses for clinical samples that had only a limited number of cells.”

Dr. Jin further explained that conventional DNase-seq provides only the average profile of DHSs using the millions of cells, giving no single-cell-specific information. With the ability to analyze DHSs in single cells, investigators will be able to better understand chromatin accessibility and gain deeper biological insight into gene expression regulation.

In their paper, the study scientists noted that scDNase-seq was both repeatable and reliable in identifying DHSs at the single-cell level. Also, they showed that the cell-specific DHSs they identified regulated cell-specific gene expression programs, highlighting the importance of understanding what is happening at the individual cell level to understand biological processes overall.

The research team applied their technique to normal and tumor cells from patients with thyroid cancer. They found thousands of tumor-specific DHSs, many of which were critical to cancer development.

“The detection and analysis of DHSs in single cells is an important step in understanding disease as well as normal cellular activity,” said Dr. Jin. “Our technique has the potential for helping researchers better understand differences in health and disease among patients with the cell-specific information it can provide. Additionally, it might assist researchers in developing targeted therapies that address specific cellular processes within individual patients.”