Shadow enhancers mediate trade-offs between transcriptional noise and fidelity.

Enhancers are stretches of regulatory DNA that bind transcription factors (TFs) and regulate the expression of a target gene. Shadow enhancers are two or more enhancers that regulate the same target gene in space and time and are associated with most animal developmental genes. These multi-enhancer systems can drive more consistent transcription than single enhancer systems. Nevertheless, it remains unclear why shadow enhancer TF binding sites are distributed across multiple enhancers rather than within a single large enhancer. Here, we use a computational approach to study systems with varying numbers of TF binding sites and enhancers. We employ chemical reaction networks with stochastic dynamics to determine the trends in transcriptional noise and fidelity, two key performance objectives of enhancers. This reveals that while additive shadow enhancers do not differ in noise and fidelity from their single enhancer counterparts, sub- and superadditive shadow enhancers have noise and fidelity trade-offs not available to single enhancers. We also use our computational approach to compare the duplication and splitting of a single enhancer as mechanisms for the generation of shadow enhancers and find that the duplication of enhancers can decrease noise and increase fidelity, although at the metabolic cost of increased RNA production. A saturation mechanism for enhancer interactions similarly improves on both of these metrics. Taken together, this work highlights that shadow enhancer systems may exist for several reasons: genetic drift or the tuning of key functions of enhancers, including transcription fidelity, noise and output.

Non-cooperative mechanism for bounded and ultrasensitive chromatin remodeling.

Chromatin remodeling is an essential form of gene regulation that is involved in a variety of biological processes. We develop a theoretical model that takes advantage of percolation effects at the level of nucleosome interactions, which allows for ultrasensitive chromatin expansion. This model is non-cooperative and readily provides spatial bounds to the expansion region, preventing uncontrolled remodeling events. We explore different chromatin architectures and the ultrasensitivity of the chromatin density as a function of transcription factor concentration. We also compare our model with experimental data involving an inhibitor of nucleosome acetylation. These results suggest a novel mechanism for spatially-bounded chromatin remodeling and they provide means for quantitative comparisons between proposed models of chromatin architecture.

Shadow enhancers can suppress input transcription factor noise through distinct regulatory logic.

Shadow enhancers, groups of seemingly redundant enhancers, are found in a wide range of organisms and are critical for robust developmental patterning. However, their mechanism of action is unknown. We hypothesized that shadow enhancers drive consistent expression levels by buffering upstream noise through a separation of transcription factor (TF) inputs at the individual enhancers. By measuring the transcriptional dynamics of several Kruppel shadow enhancer configurations in live Drosophila embryos, we showed that individual member enhancers act largely independently. We found that TF fluctuations are an appreciable source of noise that the shadow enhancer pair can better buffer than duplicated enhancers. The shadow enhancer pair is also uniquely able to maintain low levels of expression noise across a wide range of temperatures. A stochastic model demonstrated the separation of TF inputs is sufficient to explain these findings. Our results suggest the widespread use of shadow enhancers is partially due to their noise suppressing ability.

In all higher organisms, life begins with a single cell. During the early stages of development, this single cell grows and divides multiple times to develop into the many different kinds of cells that make up an organism. This is a highly regulated process during which cells receive instructions telling them what kind of cell to become. These instructions are relayed via genes, and a particular combination of activated genes determines the cell's fate. Specific pieces of DNA, known as enhancers, act as switches that control when and where genes are active, while so-called shadow enhancers are found in groups and work together to turn on the same gene in a similar way. Shadow enhancers are often active during the early stages of life to direct the formation of specialized cells in different parts of the body. But so far, it has been unclear why it is beneficial to the divide the role of activating genes across several shadow enhancers rather than a single one. Here, Waymack et al. examined shadow enhancers around a gene called Kruppel in embryos of the fruit fly Drosophila melanogaster. Manipulating the shadow enhancers showed that they help to make gene activity more resistant to changes. Factors such as fluctuations in temperature have different effects on each shadow enhancer. Having several shadow enhancers working together ensures that, whatever happens, the right genes still get activated. For genes like Kruppel, which are key for healthy development, the ability to withstand unexpected changes is a valuable evolutionary benefit. The study of Waymack et al. reveals why shadow enhancers are involved in the regulation of many genes, which may help to better understand developmental defects. Many conditions caused by such defects are influenced by both genetics and the environment. Genetic illnesses can vary in severity, which may be related to the roles of shadow enhancers. As such, studying shadow enhancers could lead to new approaches for treating genetic diseases.