Using synthetic biology to understand development

A synthetic mutual inhibition circuit interprets morphogen gradients to make sharp boundaries.

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Developmental biology is among the older disciplines of experimental biology with insights as to how organisms build themselves discovered long before we had any idea about the biomolecules that make up those organisms (Spemann and Mangold, 1923).  Enormous progress has since been made in discovering the molecular mechanisms for these developmental processes.  In addition, development has been a field in which theory and mathematical modelling have been particularly fruitful.  Thinkers from disparate backgrounds such as Alan Turing and Lewis Wolpert have conceived simple and elegant theories about how patterns can form within developing embryos and can lead to the structures we see in adult organisms.  The problem, from the perspective of the theorist, is that while their elegant mathematical constructs can replicate phenomena, they cannot prove that theirs is the mechanism at work. Conversely, the experimentalist can tease out the specifics of a particular mechanism in a particular organismal context but is often unsure of the generality of what they have found.

This is where one of the newest disciplines, synthetic biology, barges in with the hubris of the newcomer, to offer a solution.  By building synthetic genetic circuits from scratch we can test whether hypothesized mechanisms of development are truly sufficient to account for the phenomena to which they are ascribed.  Not only that but we can gain quantitative understanding of the conditions required for such phenomena to occur.  This adds to our knowledge of the basic science but also provides a framework for engineering multicellular pattern-forming systems which can pave the way for tissue engineering and self-organizing, self-repairing materials.

In our recently published paper in Nature Communications, the mechanism we have chosen to investigate using this technique is mutual inhibition downstream of morphogen induction.  This network motif has been observed in multiple developmental contexts and has been proposed as the mechanism for creating sharp boundaries (Briscoe and Small, 2015).  For our morphogens we use bacterial quorum-sensing molecules and our synthetic developmental system is composed of E. coli colonies growing on grids printed with hydrophobic ink on filter paper (Grant et al., 2016).  Building this mutual inhibition circuit required many iterations of the design-build-test-learn cycle, mainly concerned with tuning the expression level and degradation of the repressors.  Once we got the balance right, we saw this:

With one morphogen diffusing from the bottom and another diffusing from the left, there was a sharp diagonal line separating the two domains of gene expression.  This was the case even though the morphogen concentrations were changing in space throughout the whole experiment, eventually diffusing to homogeneity.  That led us to investigate the exact conditions that produce such sharp boundaries and all the experiments we performed in our paper which you can read here.

Paul Grant

Synthetic Biologist, Microsoft Research

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