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Human Chromosomes

A team of scientists at Harvard University has developed a new imaging technology for visualizing organization of chromatin, a substance within a chromosome consisting of DNA and protein, across multiple scales in single cells with high genomic throughput.

Su et al. report a multiplexed error-robust fluorescence in situ hybridization (MERFISH)-based method for genome-scale chromatin tracing and demonstrate simultaneous imaging of more than 1,000 genomic loci and nascent transcripts of more than 1,000 genes together with landmark nuclear structures. Image credit: Su et al., doi: 10.1016/j.cell.2020.07.032.

Su et al. report a multiplexed error-robust fluorescence in situ hybridization (MERFISH)-based method for genome-scale chromatin tracing and demonstrate simultaneous imaging of more than 1,000 genomic loci and nascent transcripts of more than 1,000 genes together with landmark nuclear structures. Image credit: Su et al., doi: 10.1016/j.cell.2020.07.032.

“It’s quite important to determine the 3D organization to understand the molecular mechanisms underlying the organization and to also understand how this organization regulates genome function,” said senior author Professor Xiaowei Zhuang, a researcher in the Howard Hughes Medical Institute, the Department of Chemistry and Chemical Biology, and the Department of Physics at Harvard University.

With their new imaging method, Professor Zhuang and colleagues started to build a chromosomal map from both wide-lens images of all 46 chromosomes and close-ups of one section of one chromosome.

To image something that’s still too small to image, they captured connected dots — genomic loci — along each DNA chain.

By connecting a lot of dots, they could form a comprehensive picture of the chromatin structure.

“But there was a snag. Previously, the number of dots we could image and identify was limited by the number of colors they could image together: three. Three dots can’t make a comprehensive picture,” Professor Zhuang noted.

So, the researchers came up with a sequential approach: image three different loci, quench the signal, and then image another three in rapid succession. With that technique, each dot gets two identifying marks: color and image round.

“Now we actually have 60 loci simultaneously imaged and localized and, importantly, identified,” Professor Zhuang said.

Still, to cover the whole genome, the authors needed more — thousands — so they turned to a language that’s already used to organize and store huge amounts of information: binary.

By imprinting binary barcodes on different chromatin loci, they could image far more loci and decode their identities later. For example, a molecule imaged in round one but not round two gets a barcode starting with 10.

With 20-bit barcodes, the team could differentiate 2,000 molecules in just 20 rounds of imaging.

“In this combinatorial way, we can increase the number of molecules that are imaged and identified much more rapidly,” Professor Zhuang said.

With this technique, the team imaged about 2,000 chromatin loci per cell, a more than ten-fold increase from their previous work and enough to form a high-resolution image of what the structure of chromosomes looks like in its native habitat.

They also imaged transcription activity — when RNA replicates genetic material from DNA — and nuclear structures like nuclear speckles and nucleoli.

With their high-resolution images, Professor Zhuang and co-authors determined that areas with lots of genes tend to flock to similar areas on any chromosome. But areas with few genes only come together if they share the same chromosome.

One theory is that gene-rich areas, which are active sites for gene transcription, come together like a factory to enable more efficient production.

While more research is needed before confirming this theory, one thing is now certain: local chromatin environment impacts transcription activity. Structure does influence function.

The team also discovered that no two chromosomes look the same, even in cells that are otherwise identical.

To discover what each chromosome looks like in every cell in the human body will take far more work than one lab can take on alone.

“It’s not going to be possible to build just on our work. We need to build on many, many labs’ work in order to have a comprehensive understanding,” Professor Zhuang said.

The team’s results were published in the journal Cell.

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Jun-Han Su et al. Genome-Scale Imaging of the 3D Organization and Transcriptional Activity of Chromatin. Cell, published online August 20, 2020; doi: 10.1016/j.cell.2020.07.032

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