Nanobody-tethered transposition enables multifactorial chromatin profiling at single-cell resolution

Area of Science:

• Epigenetics and Genomics
• Single-cell Biology
• Immunology

Background:

• Chromatin states are crucial for cellular function, determined by histone modifications, transcription factor binding, and DNA accessibility.
• Existing methods lack the ability to simultaneously assess multiple chromatin aspects at single-cell resolution.
• There is a need for advanced techniques to comprehensively map chromatin states in individual cells.

Purpose of the Study:

• To introduce and validate nanobody-tethered transposition followed by sequencing (NTT-seq) as a novel assay.
• To enable simultaneous, genome-wide measurement of multiple chromatin features (histone modifications, protein-DNA binding) at single-cell resolution.
• To apply NTT-seq for multimodal chromatin mapping in cell culture and human immune cells, and integrate with cell surface protein profiling.

Main Methods:

• Development of nanobody-tethered Tn5 transposase (nb-Tn5) fusion proteins.
• Utilizing a mixture of primary antibodies targeting different epitopes for multiplexed antibody binding.
• Application of NTT-seq in both bulk and single-cell formats for multimodal chromatin profiling and cell surface marker analysis.

Main Results:

• NTT-seq successfully measures up to three histone modifications and protein-DNA binding sites simultaneously at single-cell resolution.
• High-resolution multimodal maps of chromatin states were generated for cell culture and human immune cells.
• Integrated profiling of cell surface proteins and multimodal chromatin states was achieved, facilitating immune system studies.

Conclusions:

• NTT-seq is a powerful new technology for high-resolution, multimodal chromatin state analysis at the single-cell level.
• This assay overcomes limitations of previous methods, enabling a more comprehensive understanding of epigenetic regulation.
• The ability to integrate chromatin state with cell surface markers opens new avenues for studying complex biological systems like the immune system.