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Scientists at Weill Cornell Medicine and the New York Genome Center have developed a new method that maps, in single cells, the DNA binding sites of transcription factors and other regulatory proteins that control gene activity. The technique, called D&D-seq, addresses significant drawbacks of existing tools and is the first of its kind that can be readily incorporated into high-throughput, single-cell multi-omics workflows. The study was published in Cell.
Transcription factors and other DNA-binding proteins play a critical role in switching genes on and off, and a large proportion of disease-risk hotspots identified in genetics studies lie at transcription factor binding sites. Despite their importance, tools for mapping actual transcription factor binding events in cells have had notable limitations—including insensitivity to weak or transient binding events and incompatibility with standard multi-omics platforms, making it difficult to get a complete picture of how genes and gene networks are regulated.
D&D-seq— short for docking and deaminase sequencing—works by using antibodies to bring a DNA-editing enzyme close to a target protein. Even a brief interaction between the deaminase-linked protein and DNA leaves a detectable mark in sequencing data. "DNA is a marvelous molecule for recording and storing information, and we are exploiting this property to our advantage," said Ivan Raimondi, co-senior author on the study.
The team demonstrated D&D-seq by mapping binding sites of several transcription factors and chromatin remodeling proteins—proteins that influence gene activity by opening or closing the local structure of DNA. One demonstration mapped the binding sites of a key transcription factor in blood cells, comparing cells with and without a common leukemia mutation, revealing in detail how the mutation alters transcription factor binding.
Critically, D&D-seq is compatible with existing single-cell multi-omics platforms, allowing protein-DNA interaction data to be collected alongside gene activity patterns, genome sequences, and other omics layers in the same experiment. "D&D-seq is platform-agnostic—it's basically a plug-and-play feature that you can add to existing platforms to get more information from your experiments," Dr. Raimondi said.
"A lot of research has been held back because we didn't have the right tools for mapping DNA-protein interactions in single cells; and now that we have such a tool there is enormous excitement—it's really a foundational technological advance," said co-senior author Dan Landau.
Transcription factors and other DNA-binding proteins play a critical role in switching genes on and off, and a large proportion of disease-risk hotspots identified in genetics studies lie at transcription factor binding sites. Despite their importance, tools for mapping actual transcription factor binding events in cells have had notable limitations—including insensitivity to weak or transient binding events and incompatibility with standard multi-omics platforms, making it difficult to get a complete picture of how genes and gene networks are regulated.
D&D-seq— short for docking and deaminase sequencing—works by using antibodies to bring a DNA-editing enzyme close to a target protein. Even a brief interaction between the deaminase-linked protein and DNA leaves a detectable mark in sequencing data. "DNA is a marvelous molecule for recording and storing information, and we are exploiting this property to our advantage," said Ivan Raimondi, co-senior author on the study.
The team demonstrated D&D-seq by mapping binding sites of several transcription factors and chromatin remodeling proteins—proteins that influence gene activity by opening or closing the local structure of DNA. One demonstration mapped the binding sites of a key transcription factor in blood cells, comparing cells with and without a common leukemia mutation, revealing in detail how the mutation alters transcription factor binding.
Critically, D&D-seq is compatible with existing single-cell multi-omics platforms, allowing protein-DNA interaction data to be collected alongside gene activity patterns, genome sequences, and other omics layers in the same experiment. "D&D-seq is platform-agnostic—it's basically a plug-and-play feature that you can add to existing platforms to get more information from your experiments," Dr. Raimondi said.
"A lot of research has been held back because we didn't have the right tools for mapping DNA-protein interactions in single cells; and now that we have such a tool there is enormous excitement—it's really a foundational technological advance," said co-senior author Dan Landau.