New Genetic Tool Reveals How Multiple Signals Combine to Activate Genes

The human genome contains more than 20,000 genes, but in any given cell at any given moment, only a fraction of them are active. Researchers in Arnaud Krebs’s group at EMBL Heidelberg have designed a new experimental approach to study how cells decide which genes to switch on and when.

When a gene is active, its information is copied into RNA, which then guides the ribosome, the cell’s protein-building machinery. Certain DNA sequences called enhancers help control this process, since transcription factor proteins that bind them raise the odds that the associated gene becomes active. Studying subtle variations in enhancer sequences, the team found that multiple transcription factors binding together make a gene far more accessible to the transcription machinery than a single transcription factor binding alone.

A challenge in this work is separating a DNA sequence’s effects from those of its surrounding genomic environment, since every stretch of DNA carries its own epigenetic marks. To address this, the team built a genetic tool called mCHIRA, which inserts a synthetic section at a fixed genomic locus so researchers can test different regulatory sequences there one at a time. “Basically, this method enables us to insert, in a specific locus in the genome, hundreds or thousands of regulatory elements, such as enhancers or promoters, so that we could study how the genomic environment is shaping transcription factor binding, enhancer accessibility, and finally transcription,” explained Valentina Baderna, co-first author of the study published in Nature Genetics.

The work relied on single-molecule footprinting, which tracks DNA sequence and epigenetic variation across individual cells, analyzed with FootprintCharter, a computational framework built by co-first author Guido Barzaghi. “Measuring multiple aspects of DNA regulation at once, single-molecule footprinting makes for an excitingly data-rich technology,” said Barzaghi. “Tech development on both experimental and computational fronts is what allowed us to quantify molecular states at unprecedented resolution.”

“The mechanisms used by the cell to regulate gene expression are incredibly complex but follow certain basic principles,” said Krebs. “The problem is that our genomes are so complex that those principles are difficult to uncover directly. By combining synthetic biology with quantitative genomics, we can break this complexity into manageable pieces and begin to understand the rules that connect DNA sequence to gene regulation. In the future, combining these approaches with artificial intelligence will make it possible to study gene regulation at an entirely new scale.”