Everyone loves a bargain, and a new method called CoTarget really delivers. It’s a method that delivers single-cell multiomics data on multiple histone modifications and transcription factors, as well as the transcriptome. It can easily scale up to analyze a million single cells, and unlike similar methods, it doesn’t require microfluidic devices or special transposon (Tn) fusion proteins.
To get a big bang for their buck, Aibin He’s team (Peking University) developed CoTarget, which was ramped up to ultrahigh-throughput with uCoTarget. Then, adding transcriptome information led to uCoTargetX.
All of the methods are based on tagmentation. Here are the details on CoTarget, the “base” method:
- Antibodies against histone modifications or transcription factors are incubated with protein A-Tn5 assemblies that have the T7 barcoded adaptor
- Tagmentation is done sequentially with the different antibody complexes
- The second tagmentation occurs with a secondary antibody conjugated to protein A-Tn5-T5
- The T7 barcodes are all different, making it easy to pinpoint the modifications
- In a test, CoTarget simultaneously detected H3K27ac and H3K27me3 signals in human K562 (chronic myeloid leukemia) cells
To expand their offerings, the team developed uCoTarget to boost single-cell investigations to ultrahigh-throughput levels. Incorporating split-pool ligation, they could run through up to a million single cells in one experiment for even more value. Here are the details on proof-of-concept experiments with human cells:
- uCoTarget could distinguish between K562 and another human leukemia cell line
- It simultaneously detected five histone modifications
- In a comparison, uCoTarget produced similar or better results than scMulti-CUT&Tag, MulTi-Tag, and NTT-seq.
When applied to hematopoietic stem/progenitor cell (HSPC) generation, the approach also packed a punch, detecting H3K27me3, H3K27ac, and H3K4me3 in human embryonic stem cells in day 8 of a differentiation protocol. Some other interesting data:
- Based on H3K27ac info, the team could tell mesoderm, hematopoietic cells, and cardiac cells apart
- As expected, mesoderm has signals for H3K4me3 and H3K27ac around SMAD3, and for H3K27me around TAL1, important regulators of mesoderm and blood development
- Computational methods showed that cell-type enhancer binding was likely
- RUNX1, a transcription factor that promotes blood development, is enriched alongside H3K27ac at GFI1B, and is at lower levels around repressive H3K27me3 at GFI1…Both of these target genes are involved in blood development
- During HSPC development, RUNX1 binds specific enhancer regions, priming them for H3K27ac activation
But wait—there’s more! Combining uCoTarget with a version of Smart-seq2 led to uCoTargetX for chromatin and mRNA information. With the method, they could distinguish H1 (human embryonic stem cells) from K562 cells. In experiments, they saw differential enrichment of H3K27ac and RUNX1 around a particular gene, but no effects were seen in the transcriptome. This suggests that the epigenome and the transcriptome change at different times.
Overall, the CoTarget methods are a steal at a reagent cost of about $0.01 per cell, and up to five histone or transcription factor modifications plus the transcriptome can be assayed all at the same time.
See if this deal is for you at Science Advances, January 2024.