Scientists Begin To Unravel the Complexity of DNA Enhancers

Animal diversity and evolution are driven by changes in how our genetic code is expressed. Specific DNA sequences called enhancers control where, when and how strongly genes are expressed during development to create the respective organism. Studying enhancers and how they result in different patterns of gene expression therefore helps us to understand more about how evolution takes place. In addition to driving the evolution of species, enhancers are also relevant to disease: mutations in enhancers are associated with over 80% of all human diseases.

“What we see in terms of biodiversity in nature is caused, to a large degree, by changes in enhancers,” explains Justin Crocker, group leader at EMBL Heidelberg. “Understanding – and subsequently trying to predict – evolution in the time of climate change, where many animals are under the pressure to adapt quickly to fast changing environments, is an important task.”

Despite broad relevance to evolution and disease, researchers still struggle to understand how enhancers are coded in our genomes and how easy it is to reprogram them, for example to prevent or treat diseases. In an attempt to learn more about enhancers, the Crocker group from EMBL Heidelberg performed an extensive study, published in Nature, on a specific enhancer in the model organism Drosophila melanogaster, a species of fruit fly. The group discovered that this enhancer – which controls the patterns where hair grows on flies – contains a lot more information than expected.

“Whenever we changed a single letter of the enhancer DNA sequence, we created a significant change to the pattern of gene expression it drove,” explains Timothy Fuqua, PhD student at EMBL and first author of the paper. “We also found that almost all mutations to the enhancer alter the gene expression pattern in multiple ways. For example, one mutation controls not only where the expression pattern is within the fly, but also when, and how much of the gene was expressed.”

This 3D video of a fruit fly embryo was taken using the confocal microscope imaging pipeline that the Crocker group and the Advanced Light Microscopy Facility at EMBL Heidelberg developed. Green marks cells that will produce hairs and magenta marks the pattern that is controlled by a specific enhancer sequence the group studied. Credit: Timothy Fuqua/EMBL.

These results were surprising and contradict what had previously been known about enhancers. Researchers thought that these complex gene expression patterns were created by different proteins attaching to the enhancer. A first clue that this might not be true came when Crocker and his team discovered that artificially-produced enhancers did not work as designed. Their most recent results provide support for this idea. “The results showed that developmental enhancers encode a much higher level of information than previously appreciated,” Crocker says. “When we received the data, I was honestly shocked! I couldn’t believe it and we repeated everything, as we assumed that there has been a mistake.”

Importantly, the density of information encoded within the enhancer also constrains how animals can evolve. The study also showed that each possible mutation has a certain possibility for happening. This gives scientists insights into where evolution could lead. “We can use this information to predict patterns in wild fruit flies. Something which has been incredibly difficult to do so far,” Fuqua says. “Our results should encourage the community to reassess our assumption about how these regions contribute to human health.”

While studying enhancers is a well-established field in molecular biology, this study is unique in the sheer number of mutations having been studied. The group created more than 700 unique, randomly generated mutations within a single enhancer. “Nobody ever has studied so many enhancer variants at this level of depth before. It was as if evolution was happening before our very eyes!” highlights Fuqua. To perform so many experiments, the team built – assisted by the Janelia Research Campus and the Advanced Light Microscopy Facility at EMBL – a robot to handle the fly embryos used in the study, and an automated microscope pipeline to take images of each mutated line.

“Our study shows that what we have known about enhancers was oversimplified. It shows we have to study enhancers at much greater detail than ever before,” Fuqua says. Therefore, in the next step, the team not only wants to expand the pipeline and its throughput, but also plans to study other enhancers and see if they can observe similar effects. “Can what we found be applied to other enhancers or not? We don’t know yet. But we plan to find out,” concludes Crocker.

Reference: Fuqua T, Jordan J, van Breugel ME, et al. Dense and pleiotropic regulatory information in a developmental enhancer. Nature. 2020. doi:10.1038/s41586-020-2816-5.

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